US 2007.0122492A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2007/0122492 A1 Behr et al. (43) Pub. Date: May 31, 2007

(54) PLANT EXTRACTS AND Publication Classification DERMATOLOGICAL USES THEREOF (51) Int. Cl. A6IR 36/45 (2006.01) A6IR 36/53 (2006.01) A 6LX 36/899 (2006.01) (76) Inventors: Stephen Behr, Outremont (CA); A6IR 36/8962 (2006.01) Philippe Duret, Quebec (CA); Nathalie A6IR 36/906 (2006.01) Gendron, Neuville (CA); Johane A6IR 36/48 (2006.01) Guay, St-Augustin de Desmaures (CA); A6IR 36/8 (2006.01) Bernard Lavallee, Ste. Foy (CA); A6IR 36/3 (2006.01) Brigitte Page, Quebec (CA) A6IR 36/23 (2006.01) (52) U.S. Cl...... 424/725; 424/750; 424/754; 424/732; 424/760; 424/770; 424/757; 424/756; 424/745 Correspondence Address: HELLEREHRMAN LLP (57) ABSTRACT 275 MODLEFIELD ROAD MENLO PARK, CA 94025-3506 (US) The present invention provides for plant extracts and der matological formulations comprising one or more plant extracts that are capable of inhibiting one or more extracel lular proteases selected from the group of matrix metallo protease-1 (MMP-1), matrix metalloprotease-2 (MMP-2), (21) Appl. No.: 10/533,025 matrix metalloprotease-3 (MMP-3), matrix metallopro tease-9 (MMP-9) and human leukocyte elastase (HLE). The present invention further provides for a rapid method for screening plant extracts to identify those having the above (22) PCT Fed: Nov. 18, 2004 activity that are suitable for incorporation into the derma tological formulations of the invention. The invention also provides for the use of the plant extracts as dermatological agents Suitable for the treatment or prevention of various (86) PCT No.: dermatological conditions, including wrinkling or sagging of the skin, irradiation induced skin and/or hair damage, S 371(c)(1), deepening of skin lines, elastotic changes in the skin, as well (2), (4) Date: Nov. 20, 2006 as for the routine care of the skin, hair and/or nails. Patent Application Publication May 31, 2007 Sheet 1 of 9 US 2007/O122492 A1 FIGURE 1

Potential Plant

Optional Pre-Harvest Treatment

Harvest Solid Sl

Optional Storage Treatment

us an X is 88 as a Contact Solid S1 with Solvent A Extraction Process I Separate Extract A m from Solid Matter S2 Potential Extract A

Extraction Process II Separate Extract B from Solid Matter S3

Contact Solid S3 with Solvent C Extraction Process III

Separate Extract C from Solid Matter S4

Solid S4 Patent Application Publication May 31, 2007 Sheet 2 of 9 US 2007/0122492 A1 FIGURE 2

Potential Extract

Optional Separation Procedure(s)

Potential Extract

Test Aliquots against OC O. O. Extracellular Proteases

Does Extract demonstrate inhibitory Potential Extract is an Inconclusive Extract of the activity against one or more extracellular Invention

proteases?

Potential Extract is not an Extract of the Invention

Plant remains a Potential Plant

Entire Procedure may be repeated on Potential Plant under varied conditions Patent Application Publication May 31, 2007 Sheet 3 of 9 US 2007/0122492 A1 FIGURE 3

One or More Plants of the Invention

Optional Pre-Harvest Treatment

Harvest Solid Sl

Optional Storage Treatment

Extraction Process I, II or II. Process may be sequentially repeated

Purification Procedure(s)

Quality Control

Extract Demonstrating Inhibitory Activity Against One or More Extracellular Proteases

Prepare Formulation for Use Patent Application Publication May 31, 2007 Sheet 4 of 9 US 2007/0122492 A1 FIGURE 4

Patent Application Publication May 31, 2007 Sheet 5 of 9 US 2007/O122492 A1

FIGURE 5 Patent Application Publication May 31, 2007 Sheet 6 of 9 US 2007/O122492 A1

FIGURE 6

Dried powdered material Analytical Scale: 2 - 10 g Large scale: 2-3 kg

Analytical scale: procedure 1 Large scale: procedure 2

Centrifugation

EtOHIMeoH extract EtOHIwater extract

Concentration under reduce pressure EtOHIMeOH extract

Dilution with 10-5% water

Partition with hexane

Extract B Hexane extract C

1) Concentration Procedure 2) Solubilization EtOH water 20/80 viv EtOHIMeOh:8515 why 3) Partition with Ethyl acetate 2x100ml protocola) SOHNWater 85/15 wiv 2x100ml (protocol b) Blender (2 x 10 min) Room temperature Procedure 2: Evaporation to dryness OHIMech:85/15 viv (protocola) EtOHWatar: 85/15 yN Aqueous extract E ACOEt extract F maceration(protocol b) percolation A. Room terrperattire Patent Application Publication May 31, 2007 Sheet 7 of 9 US 2007/0122492 A1

— Melilotus alba -e-keratinocytes

-O-Fibroblasts

80

6 O

-e-keratinocytes -O- Fibroblasts

FIGURE 7 Patent Application Publication May 31, 2007 Sheet 8 of 9 US 2007/0122492 A1

1009wÐ (OS:OS)OZH:ng»Dow (08:0Z)OZH:ngXpow (0:001)ozH:ng»Dow

FIGURE 8 Patent Application Publication May 31, 2007 Sheet 9 of 9 US 2007/0122492 A1

a 3S. SE. 5E. 5E 5E 5E saE 9 EN S.E on s in o un o o g 2 2 3

ac U nO y ON Ms w wrd R.o N ra a.c -- O cs d c ES o O : E isd C ro cn cs 5 S.E <: S5 s Cd o ?u Vs t cyL c) - Q C > d- ? s - O o 3 U O U g s s s o . --a S.d — 3 is E is

FIGURE 9 US 2007/01 22492 A1 May 31, 2007

PLANT EXTRACTS AND DERMATOLOGICAL activity of MMP-1, -2, -3 and -9 in the skin has also been USES THEREOF shown to occur in response to extrinsic factors such as UV exposure (see U.S. Pat. No. 5,837.224). The ageing process FIELD OF INVENTION (both chronological and photo-induced) involves the increased breakdown various components of the ECM in the 0001. The invention pertains to the field of dermatology, skin, notably collagen, elastin and fibronectin. Enhanced specifically within the field of dermatological preparations expression of collagenase (MMP-1) and stromelysin-1 comprising plant extracts. (MMP-3) has been described as playing a central role in connective tissue breakdown in the skin (Brenneisen, et al., BACKGROUND OF THE INVENTION (2002) Ann. N.Y. Acad. Sci., 973:31-43). Similarly, increased expression of serine elastase in hairless mouse 0002 The skin is the most environmentally-stressed models of chronological and photo-ageing was shown to organ in mammals, particularly in humans. Not only is the skin Subjected to germs, toxic chemicals and hostile envi result in increased fibronectin degradation (Labat-Robert, et ronments, it is the only organ directly exposed to ultraviolet al., (2000) J Photochem. Photobiol. B., 57:113-118). light (UV). In addition, the vitality of this organ is a 0005 Elastic fibers are essential extracellular matrix consequence of genetic processes, which over time, lead to macromolecules comprising an elastin core surrounded by a a decrease in the functionality of the skin. Hence, a variety mantle of fibrillin-rich microfibrils. These fibers endow of dermatological conditions may occur as a result of connective tissues such as blood vessels, lungs and skin with ongoing intrinsic factors (for example, chronological age the critical properties of elasticity and resilience (see review ing, disease and allergies) and/or exposure to a number of of elastic fibers by Kielty C M et al: J Cell Sci (2002) extrinsic factors (such as infection, trauma, radiation, toxins 115:2817-2828). Exposure to the sun is known to cause and steroid use). Skin is a highly organized structure con disorganization of elastin in the skin known as "elastosis.” sisting of two principal parts. The outer thinner part, the which is also a hallmark of skin-ageing. Neutrophil elastase epidermis or cuticle, is organised into four or five cell layers has been implicated in elastosis, for example, when com depending on its location. These layers are the stratum pared to normal mice, mice that are deficient in neutrophil corneum, stratum lucidem (usually only present where the elastase are unaffected by exposure to UVB. In addition, an skin is thickened), stratum granulosum, stratum spinosum increase in elastase activity has been observed in the skin and stratum basale. The inner, thicker part of the skin, the following chronic UVB irradiation (Tsukahara Ketal Biol dermis or true skin, is composed of a papillary layer above Pharm Bull 2001:24(9):998-1003). Both a synthetic inhibi and a reticular layer below. The dermis also comprises blood tor of fibroblast elastase and an extract of Sanguisorba vessels, nerves, hair follicles and Sweat glands. The layer officinalis L. inhibited wrinkle formation and maintained below the dermis, the hypodermis, comprises mainly loose skin elasticity in the rat (Tsukahara Ketal Biol Pharm Bull connective tissue and adipose cells and may be considered 2001:24(9):998-1003). part of the skin in that it functions to anchor the epidermis/ 0006 MMPs also play a role in the loss of elastic fibers dermis to the underlying bone and muscle. The hypodermis in skin. Tissue loss during ageing and age-dependent also supplies the dermis with blood vessels and nerves. pathologies are the result of a disturbed regulation of pro 0003. The cells of the skin, like many tissues, are gen teolytic activities in which elastase-type endopeptidases, erally in contact with a network of large extracellular especially MMP-2 and -9, are overactivated (Isnard Netal: macromolecules that occupy the spaces in a tissue between Biomed Pharmacother. 2002 July:56(5):258-64). In addi the component cells and between adjacent tissues. This tion, gelatinase B (MMP-9) has been shown to degrade extracellular matrix (ECM) functions as a scaffolding on fibrillin in human skin tissue sections (Berton Aetal, Matrix which the cells and tissue are supported and is involved Biol 2000:19(2): 139-148). actively in regulating interaction of the cells that contact it. 0007. In an effort to ameliorate the vast number of The principal macromolecules of the ECM include the dermatological disorders, treatments spanning topical collagens (the most abundant proteins in the body) and therapy (creams, oils, lotions, gels and sprays) to oral glycosaminoglycans (complex polysaccharides which are therapy, cosmetic procedures, injections and ultraviolet usually bonded to protein and then termed proteoglycans). therapy have been developed. Topical skin applications, for Additional proteins that may be found in the ECM include example; are known in the art to help shield the skin from elastin, fibronectin and laminin. The dermal layer of the skin the Vagaries of the environment. Conventional skin protec is composed largely of ECM (or “connective tissue') con tions typically attempt to either protect the skin from UV taining high proportions of collagen and elastin in which light (see U.S. Pat. No. 5,141,741) or provide additional cells are embedded. agents capable of neutralizing free radicals (U.S. Pat. No. 0004 Components of the ECM are degraded by extra 6,764,693). Methods of inhibiting either chronological or cellular proteolytic enzymes that are secreted locally by photo-ageing of the skin by application of UV blocking cells. Extracellular proteases, in particular matrix metallo compounds in combination with compounds that inhibit proteinases (MMPs), have been implicated in a number MMPs have also been reported (U.S. Pat. Nos. 5,837,224; dermatological conditions, for example, in both chronologi 6,130,254 and 6,365,630 and U.S. Patent Application No. cal ageing and photo-ageing processes involve extracellular 20010053347). Mercaptoketone and mercaptoalcohol com proteases (see, for example, U.S. Patent Application No. pounds that inhibit the activity of MMPs and their use in 200100513347). An age-related increase in levels of MMPs, treating or controlling disease states such as arthropathy, in particular MMP-1, -2 and -9, in the skin has been dermatological conditions, bone resorption, inflammatory demonstrated (see U.S. Patent Application No. diseases and tumor invasion have also been described (U.S. 200100513347). An analogous increase in the level and/or Pat. No. 6,307.101). Addition of certain plant extracts or US 2007/01 22492 A1 May 31, 2007

phyto-compounds to preparations, such as lotions, creams (U.S. Pat. No. 6,432,456) and from Brassica olearacea and gels, to treat dermatological disorders has also been (U.S. Pat. No. 6,177,122) have also been described. reported. These cosmetic compositions serve to shield the skin from UV light (U.S. Pat. Nos. 4,857,325; 5,141,741 and 0011. The effect of methanol extracts from medicinal 6,342,208) and act as antioxidants in the neutralization of plants on elastase activity has been reported by Lee and Kim free radicals (U.S. Pat. No. 4,923,697). Some fruit extract (Inter. J. of Cosmetic Sci. 21:71-82 (1999)). Of approxi containing dermatological agents, capable of neutralizing mately 150 extracts screened only the methanol extracts of A. catechu, C. Cassia, M. fragrams, C. longa, A. katsumadia, free radicals, additionally moisturize and facilitate the and D. Cassirrhizoma demonstrated good inhibition of hydration of the skin (see U.S. Pat. No. 6,800,292). elastase activity. Similarly, peptide-containing extracts of L. 0008. Other plant extracts useful in dermo-cosmetics albus (PCT/FR00/01007, Publication No. WO 00/62789) have been described (see U.S. Pat. Nos. 6,682,763; 5,824, have been shown to inhibit the activity of extracellular 320 and 6,406.720). Here, external agents derived from proteases including MMP-1, MMP-2 and MMP-9, using olive plants are reported as having skin-beautifying effects, fibroblast models. in particular, an anti-ageing effect related to the prevention and elimination of wrinkles and sags of the skin (U.S. Pat. 0012 A process for obtaining plant extracts capable of No. 6,682.763). Furthermore, a whitening effect, which can inhibiting various extracellular proteases has been described lighten (U.S. Pat. No. 5,073.545) or prevent dark skin, in International Patent Application PCT/CA02/00285 (Pub melasma, ephelis and darkening or dullness of the skin has lication No. WO 02/06992), in which the extracts were been reported (U.S. Pat. No. 6,682,763). Dermo-cosmetics screened on the basis of their ability to inhibit extracellular containing plant extracts for application to the mucous proteases in in vitro assays. The ability of these extracts to membrane or exoskeleton, in addition to the skin, have also inhibit extracellular proteases in vivo or to inhibit processes been considered (U.S. Pat. No. 6,406.720); the active ingre associated with the activity of Such proteases, however, was dient of these cosmetics being derived from Spondias mom not described or Suggested. bin, Maproumea guianensis, Waltheria indica, Gouania 0013 This background information is provided for the blanchetiana, Cordia Schomburgkii, Randia armata or purpose of making known information believed by the Hibiscus fircellatus; Plant extracts useful in the treatment of applicant to be of possible relevance to the present inven eczema and/or psoriasis (U.S. Pat. Nos. 6,676.975 and tion. No admission is necessarily intended, nor should be 4,855,131), hemorrhoids (U.S. Pat. No. 5,627,216) and for construed, that any of the preceding information constitutes maintaining general skin care (U.S. Pat. No. 6,193.975) have prior art against the present invention. also been described. 0009. A number of patents and publications report the SUMMARY OF THE INVENTION inhibition of one or more extracellular proteases by com 0014) An object of the invention is to provide plant pounds extracted from plants. For example, Sun et al., extracts and dermatological uses thereof. In accordance with (1996) Phytotherapy Res., 10: 194-197, reports the inhibi one aspect of the present invention, there is provided a tion in vitro of stromelysin (MMP-3) and collagenase by dermatological formulation comprising a physiologically betulinic acid extracted from Doliocarpus verruculosis. acceptable carrier and an effective amount of one or more SaZuka et al. (1997) Biosci. Biotechnol. Biochem., 6.1: plant extracts having extracellular protease inhibiting activ 1504-1506, reports the inhibition of gelatinases (MMP-2 ity, said plant extract derived from any one of the plants and MMP-9) and metastasis by compounds isolated from listed in Tables 1, 2, 3, 4 and 5 by solvent extraction, said green and black teas. Kumagai et al., JP 08104628 A2, Apr. extracellular protease selected from the group of matrix 1, 1996 (CA 125: 67741) reports the use of flavones and metalloprotease-1 (MMP-1), matrix metalloprotease-2 anthocyanines isolated from Scutellaris baicanlensis roots (MMP-2), matrix metalloprotease-3 (MMP-3), matrix met to inhibit collagenase. Gervasi et al., (1996) Biochem. Bio alloprotease-9 (MMP-9) and human leukocyte elastase phys. Res. Comm., 228: 530-538, reports the regulation of (HLE), wherein said extract affects one or more cellular MMP-2 by some plant lectins and other saccharides. Dubois activities in skin cells. et al., (1998) FEBS Lett., 427:275-278, reports the increased 0015. In accordance with another aspect of the present secretion of deleterious gelatinase-B (MMP-9) by some invention, there is provided a plant extract having extracel plant lectins. Nagase et al., (1998) Planta Med., 64: 216 lular protease inhibiting activity, said plant extract derived 219, reports the weak inhibition of collagenase by delphini by solvent extraction from a plant selected from the group din, a flavonoid isolated from Solanum melongena. of Aconitum napellus, Acorus calamus, Alchemilla mollis, 0010. Other reports include Asano et al. (1998) Immu Allium cepa, Allium sativum, Allium tuberosum, Ambrosia nopharmacology, 39: 117-126), which describes the inhibi artemisuifolia, Anethum graveolens, Anthemis tinctoria, tion of TNF-C. production using Tripterygium wilfordii Hook Aronia melanocarpa (Michx.) Ell. Arctostaphylos uva-ursi, F. extracts; Maheu et al. (1998) Arthritis Rheumatol, 41: Aroniaxprunifolia, Artemisia dracunculus, Avena sativa, 81-91), which reports the use of avocado/soy bean non Beta vulgaris, Beta vulgaris L. Subsp. Vulgaris, Borago saponifiable extracts in the treatment of arthritis: Makimura officinalis, Brassica napus, Brassica oleracea, Brassica et al. (1993) J Periodontol., 64: 630-636), which reports oleracea L. var. italica Plenck, Brassica rapa, Bromus the use of green tea extracts to inhibit collagenases in vitro inermis, Capsicum annuum L. var. annuum, Cerastium and Obayashi et al. (1998) Nippon Keshonin Gijutsusha tomentosum, Chaerophyllum bulbosum, Chenopodium Kaishi, 32: 272-279 (CA 130: 92196)), which reports the quinoa, Chenopodium quinoa Subsp. Quinoa, Chenopodium inhibition of collagenase-I (MMP-1) from human fibroblast quinoa Willd. Chichorium endivia, Chichorium endivia and neutrophil elastase by plant extract from Eucalyptus and Subsp. Endivia, Circium arvense, Citrullus lanatus, Cornus Elder. Plant extracts derived from Capsicum Annuum L Canadensis, Cornus sericea, Cynara cardunculus Subsp. US 2007/01 22492 A1 May 31, 2007

Cardunculus, Daucus carota, Daucus carota Subsp carota 0023. In accordance with another aspect, there is pro L., Dolichos lablab, Euphorbia amygdaloides, Fagopyrum vided a use of a plant extract of the present invention for the tataricum, Foeniculum vulgare, Frangula alnus, Galinsoga routine care of the skin, hair and/or nails. quadriradiata, Gentiana lutea, Geranium sanguineum, Geraniumxcantabrigiense, Glycyrrhiza glabra, Hamamelis 0024. In accordance with another aspect, there is pro virginiana, Helianthus strumosus, Heliotropium arbore vided a use of a plant extract of the present invention to scens, Hordeum vulgare Subsp. Vulgare, Hypomyces lactif improve the health and/or appearance of the skin, hair and/or luorum, Juniperus communis L., Lentinus edodes, Lotus nails. corniculatus, Manihot esculenta, Matricaria recutita, 0025. In accordance with another aspect, there is pro Melilotus albus, Melilotus alba Medik, Melissa officinalis, vided a use of a plant extract of the present invention in the Menthaxpiperita, Oenothera biennis, Pastinaca sativa L., treatment or prevention of a dermatological condition. Petroselinum Crispum, Phaseolus vulgaris, Physalis phila delphica, Phytolacca decandra, Phytolacca decandra syn. 0026. In accordance with another aspect, there is pro P. americana, Pimpinella anisum, Pisum sativum, Potentilla vided a use of a plant extract of the present invention to anserina L., Potentilla fruticosa, Poterium sanguisorba, attenuate or prevent skin ageing. Pyrus communis, Raphanus raphanistrum, Rheumxhybri 0027. In accordance with another aspect of the present dum, Rhus typhina L., Ribes nigrum L., Ribes Sylvestre, invention, there is provided a process for identifying a plant Rodgersia spp., Rosmarinus officinalis, Rubus Occidentalis, extract Suitable for the preparation of a dermatological Rubus thibetanus, Rumex Crispus, Rumex scutatus, Ruta formulation, said process comprising the steps of: (a) gen graveolens, Salvia officinalis, Sambucus Canadensis L., erating a plurality of potential extracts by Solvent extraction Setaria italica, Solanum melongena L., Sorghum dochna of plant material; (b) analysing the ability of each of said bicolor gr technicum, Stellaria media, Tanacetum cinerarii potential plant extracts to inhibit one or more extracellular folium, Taraxacum officinale, Teucrium chamaedrys, Thy protease selected from the group of matrix metallopro mus fragantissimus, ThymusXcitriodorus, Trifolium incar tease-1 (MMP-1), matrix metalloprotease-2 (MMP-2), natum, Triticosecale spp., Tropaeolum maius L., Tsuga matrix metalloprotease-3 (MB-3), matrix metalloprotease-9 Canadensis, Tsuga diversifolia, Vaccinium angustifolium, (MMP-9) and human leukocyte elastase (HLE); (c) selecting Vaccinium angustifolium Ait. Vitia sp., xTriticosecale spp., those potential extracts that are capable of inhibiting the Zea mays L. and Zingiber officinale, and said extracellular activity of at least one of said extracellular proteases to protease selected from the group of matrix metallopro provide a group of extracts; (d) analysing each extract in said tease-1 (MMP-1), matrix metalloprotease-2 (MMP-2), group of extracts for the ability to affect one or more cellular matrix metalloprotease-3 (MMP-3), matrix metallopro activities in skin cells selected from the group of attenuating tease-9 (MMP-9) and human leukocyte elastase (HLE). the breakdown of collagen, fibronectin, fibrillin and/or elas 0016. In accordance with another aspect of the present tin, attenuating endothelial cell migration; increasing col invention, the plant extract is derived from a plant selected lagen production; attenuating UV-induced extracellular pro from the group of Beta vulgaris L., Brassica oleracea L., tease activity and attenuating tractional forces generated by Capsicum annuum L. Chenopodium quinoa, Daucus carota fibroblasts; and (e) selecting an extract that is capable of L., Geraniumxcantabrigiense, Juniperus communis L., affecting one or more of said cellular activities to provide a Melilotus alba, Pastinaca sativa L., Potentilla anserina L., plant extract Suitable for the preparation of a dermatological Rhus typhina L., Solanum melongena L., Tropaeolum maius formulation. L. Vaccinium angustifolium, XTriticosecale spp. and Zea BRIEF DESCRIPTION OF THE FIGURES may’s L. 0028 FIG. 1 presents an overview of a procedure that can 0017. In accordance with another aspect of the present be followed in accordance with one embodiment of the invention, the plant extract is derived from the plant material invention in order to generate plant extracts, each of which by extraction with an alcohol, water, an aqueous buffer, or a is derived from solid plant material; combination thereof as solvent. 0018. In accordance with another aspect, there is pro 0029 FIG. 2 describes in further detail, the procedure of vided a use of a plant extract of the invention in the FIG. 1; preparation of a dermatological formulation. 0030 FIG. 3 presents an overview of a commercial 0019. In accordance with another aspect, there is pro procedure that can be followed to prepare plant extracts vided a use of a dermatological formulation of the present based on the procedure of FIG. 1; invention for the routine care of the skin, hair and/or nails. 0031 FIG. 4 shows the effect of a plant extract of the 0020. In accordance with another aspect, there is pro invention derived from Rheum rhabarbarum on cord for vided a use of a dermatological formulation of the present mation, (a) untreated cells; (b) cells treated with a positive invention to improve the health and/or appearance of the control; (c) cells treated with an extract of the invention (1x skin, hair and/or nails. concentration), and (d) cells treated with an extract of the 0021. In accordance with another aspect, there is pro invention (2x concentration); vided a use of a dermatological formulation of the present 0032 FIG. 5 presents an overview of a procedure that can invention in the treatment or prevention of a dermatological be followed in another embodiment of the invention in order condition. to generate plant extracts, each of which is derived from 0022. In accordance with another aspect, there is pro Solid plant material; vided a use of a dermatological formulation of the present 0033 FIG. 6 describes in further detail, the procedure of invention to attenuate or prevent skin ageing. FIG. 5; US 2007/01 22492 A1 May 31, 2007

0034 FIG. 7 depicts the effect of plant extracts of the condition or of exposure of the skin to harmful elements invention on the viability of human keratinocytes and fibro (such as UV irradiation), or they can occur naturally, for blasts; example, as part of the ageing process. 0035 FIG. 8 depicts the effect of plant extracts of the 0040 Accordingly, inhibition of skin EPs can attenuate invention on the production of collagen in human dermal undesirable EP-mediated ECM degradation in the skin and fibroblasts; and structural changes associated therewith. One embodiment of 0036 FIG. 9 depicts the effect of plant extracts of the the present invention provides for plant extracts that are invention on the release of IL-8 from human skin kerati capable of attenuating undesirable EP-mediated ECM deg nocytes. radation in the skin and structural changes associated there with. EP-mediated ECM degradation refers to the break down of one or more component of the ECM surrounding DETAILED DESCRIPTION OF THE the cells of mammalian skin including, for example, col INVENTION lagen, elastin, fibrillin and/or fibronectin. Undesirable skin 0037. The present invention provides for dermatological structural changes include, for example, wrinkling and/or formulations comprising one or more plant extracts that are sagging of the skin, loss of elasticity, redness, inflammation, capable of inhibiting at least one skin extracellular protease formation of lesions, thinning of the epithelium, abnormal (EP). In the context of the present invention, the terms “skin migration of cells within the skin (such as that which occurs extracellular protease' and “skin EP refer to the extracel during angiogenesis or inflammation), or various combina lular proteases: matrix metallpprotease-1 (MMP-1), matrix tions thereof. metalloprotease-2 (MMP-2), matrix metalloprotease-3 (MMP-3), matrix metalloprotease-9 (MMP-9) and human 0041) Definitions leukocyte elastase (HLE). The present invention further 0042 Unless defined otherwise, all technical and scien provides for a rapid method for screening plant extracts to tific terms used herein have the same meaning as commonly identify those having the above activity that are suitable for understood by one of ordinary skill in the art to which this incorporation into the dermatological formulations of the invention belongs. invention. The invention also provides for the use of plant extracts having the above activity as dermatological agents 0043. The term “potential plants, as used herein, is suitable for the treatment or prevention of various derma intended to include all species of the Kingdom Plantae, tological conditions, including wrinkling or sagging of the including terrestrial, aquatic or other plants under the Divi sion Chlorophyta, Division Rhodophora, Division Paeo skin, irradiation-induced skin and/or hair damage, deepen phyta, Division Bryophyta and Division Tracheophyta; Sub ing of skin lines, elastotic changes in the skin, and the like, division Lycopsida, Subdivision Sphenopsida, Subdivision as well as for the routine care of the skin, hair and/or nails Pteropsida and Subdivision Spermopsida; Class Gymno and to improve the health and/or appearance of the skin, hair spermae, Class Angiospermae, Subclass Dicotyledonidae and nails. and Subclass Monocotyledonidae. 0038. The present invention additionally provides for novel plant extracts identified by the methods described 0044) The term “plant material.” as used herein, refers to herein that inhibit one or more skin extracellular proteases, any part or parts of a plant taken either individually or in a and which are suitable for use as dermatological agents. group. Examples include, but are not limited to, leaves, Semi-purified/purified active ingredients (i.e. molecules or flowers, roots, seeds, pods, stems, fruits, seed coats, buds, compounds) isolated from a plant extract of the invention and other parts of a plant. and the use of these active ingredients, alone or in combi 0045. The term “potential extract,” refers to a composi nation with an extract, as dermatological agents are also tion prepared by contacting plant material with a solvent contemplated. following the procedures described herein, which has not yet 0.039 The integumentary system of a mammal is made up been determined to possess inhibitory activity against one or of components (the skin, hair and nails) derived from the more extracellular protease. The potential extract can ectoderm and Subjacent mesoderm. Mammalian skin is optionally be subjected to one or more separation 1 and/or composed of a number of layers of cells embedded in an purification step. extracellular matrix (the ECM), which provides structure to 0046) The term “plant extract of the invention,” as used the skin and comprises a number of polymeric structural herein, refers to a composition prepared by contacting plant components including collagen, elastin and fibronectin. Dis material with a solvent following the procedures described persed within the ECM are various types of cells, including herein, which demonstrates inhibitory activity against one or fibroblasts and immune cells, which secrete EPs into the more extracellular protease selected from the group of ECM. The ECM of the skin is in a constant state of flux, or matrix metalloprotease-1 (MMP-1), matrix metallopro turnover, which is tightly regulated and mediated in part by tease-2 (MMP-2), matrix metalloprotease-3 (MMP-3). the secreted EPs, which are capable of degrading the struc matrix metalloprotease-9 (MMP-9) and human leukocyte tural components of the ECM. A shift in this turnover to an elastase (HLE). The plant extract can be a primary extract or increased rate in the breakdown of one or more ECM a Substantially pure extract. structural components, such as collagen.(s) or elastin, results in an increased degradation of the ECM and undesirable 0047 The terms “substantially purified’ and “substan structural changes in the skin itself. Changes in the structure tially pure' when used in reference to F a plant extract of the of the ECM can also affect the hair and nails, which are invention refer to an extract that has been subjected to at reliant on the skin for nourishment. Shifts in the balance of least one additional treatment Subsequent to a first solvent ECM turnover can occur as a consequence of a disease extraction of plant material. Thus the present invention US 2007/01 22492 A1 May 31, 2007 provides for primary extracts that result from a “one-step' components of the integumentary system of a subject, i.e. on solvent extraction of plant material followed optionally with the skin, hair or nails, caused by ageing or by intrinsic or a filtration or centrifugation step, and for Substantially pure extrinsic factors. plant extracts that have been Subjected to one or more additional steps, such as liquid-liquid extraction, Solid-liquid 0054 The term “treatment, as used herein, refers to an extraction, chromatography, distillation, evaporation, filtra intervention performed with the intention of improving a tion, and the like following the initial extraction process. recipient's status. The improvement can be subjective or Both primary extracts and Substantially pure extracts are objective and is related to the amelioration of the symptoms encompassed by the term “plant extracts of the present associated with, preventing the development of or altering invention.” the pathology of a condition being treated. Thus, the term treatment is used in the broadest sense, and includes the 0.048. The term “stressor,” as used herein, refers to a prevention (prophylaxis), moderation, reduction, and curing factor, such as a physical factor, a chemical compound, or a of a condition at various stages. Prevention of deterioration biological agent that is used to activate a defense response of a recipient’s status is also encompassed by the term. in a plant and thereby elicit production of various chemicals, Those in need of treatment include those already having the including extracellular protease inhibitors. Elicitors and condition as well as those prone to, or at risk of developing, inducers are also considered to be stressors. The term the condition and those in whom the condition is to be "isolated when used in reference to an active ingredient, prevented. Such as a molecule or compound, refers to a form of the active ingredient that has been removed from the plant tissue 0055. The term “ameliorate” or “amelioration” includes from which it is derived. Typically, an isolated active the arrest, prevention, decrease, or improvement in one or ingredient is relatively free of proteins, nucleic acids, lipids, more the symptoms, signs, and features of the condition carbohydrates or other materials with which it is naturally being treated, both temporary and long-term. associated in a plant. An isolated active ingredient may be further purified using routine and well-known methods such 0056. The term “subject” or “patient,” as used herein, as those described herein. As such, an isolated active ingre refers to a mammal in need of treatment or who would dient of the invention can constitute at least about one or a otherwise benefit from the use of a dermatological formu few percent of a sample, for example, at least about five lation of the invention. percent. In one embodiment, the isolated active ingredient 0057. As used herein, the term “about” refers to a constitutes at least about twenty percent of a sample. In +/-10% variation from the nominal value. It is to be another embodiment, the isolated active ingredient is further understood that such a variation is always included in any purified to constitute at least about fifty percent of a sample. given value provided herein, whether or not it is specifically In other embodiments, the isolated active ingredient can be referred to. further purified to constitute at least about eighty percent, at least about ninety percent and at least about ninety-five 0058 Other chemistry terms herein are used according to percent or more of a sample. conventional usage in the art, as exemplified by The McGraw-Hill Dictionary of Chemical Terms (ed. Parker, S., 0049. The term “skin cell,” as used herein, refers to a cell 1985), McGraw-Hill, San Francisco). normally present within the skin of a mammal. "Skin’ refers to the epidermis (including the stratum germinativum, Stra 0059) Plant Extracts tum spinosum, stratum granulosum, stratum lucidum and stratum corneum), the dermis (including the papillary der 0060. The present invention provides for plant extracts mis and the reticular dermis) and the hypodermis. The term Suitable for use as dermatological agents. In accordance with “skin cells' thus includes, but is not limited to, kerati the present invention, the plant extracts are capable of nocytes, fibroblasts, endothelial cells (including vascular inhibiting one or more skin extracellular proteases selected endothelial cells), basal cells, granular cells, Merkel cells, from the group of matrix metalloprotease-1 (MMP-1), melanocytes, Langerhans cells, leukocytes, mastocytes, matrix metalloprotease-2 (MMP-2), matrix metallopro nerve cells, adipose cells and macrophages. tease-3 (MMP-3), matrix metalloprotease-9 (MMP-9) and human leukocyte elastase (HLE). 0050. The term “attenuate,” as used herein, means to slow-down, inhibit or prevent. 0061 While some plant extracts have previously been identified that inhibit one or more extracellular proteases, 0051. The term “cell migration,” as used herein, refers to the potential for plant extracts that inhibit any one of this the movement, typically abnormal, of a cell or cells from one particular group of proteases to be effective in various locus to another. Examples of cell migration include the dermatological applications has not previously been estab movement of cells through the ECM or basal lamina during lished. The systematic evaluation of a large plant library to angiogenesis. identify extracts from the plants in this library that inhibit 0.052 A“dermatological agent,” as used herein, refers to one or more of this particular group of proteases and the an extract, compound, composition or formulation intended Subsequent evaluation of the extracts in cellular models as for the routine care of the integumentary system, for improv described herein has allowed this potential to be recognised. ing the health and/or appearance of the integumentary Accordingly, the plant extracts of the invention suitable for system or for the treatment or prevention of a dermatological use as dermatological agents inhibit at least one of the above condition. listed skin EPs. The present invention also contemplates plant extracts that inhibit two or more, three or more, four or 0053. The term “dermatological condition,” as used more, or all five of MMP-1, MMP-2, MMP-3, MMP-9 and herein, refers to a condition present on one or more of the HLE. US 2007/01 22492 A1 May 31, 2007

0062. In one embodiment of the present invention, the Tsuga diversifolia, Vaccinium angustifolium, Vaccinium plant extract is capable of inhibiting at least P-1. In another angustifolium Ait. Vitia sp., xTriticosecale spp., Zea mays embodiment, the plant extract is capable of inhibiting at least L. and Zingiber officinale. MMP-2. In a further embodiment, the plant extract is 0066. In another embodiment, the plant extracts are capable of inhibiting at least MMP-3; In another embodi derived from a plant selected from the group of Allium cepa, ment, the plant extract is capable of inhibiting at least Allium sativum, Ambrosia artemisiifolia, Ambrosia arte MMP-9. In another embodiment, the plant extract is capable misiifolia, Arctostaphylos uva-ursi, Aroniaxprunifolia, Arte of inhibiting at least HLE. misia dracunculus, Avena sativa, Beta vulgaris, Beta vul garis L. Subsp. Vulgaris, Brassica napus, Brassica oleracea, 0063. In an alternative embodiment, the plant extract is Brassica oleracea L. var. italica Plenck, Brassica rapa, capable of inhibiting at least two of MMP-1, MMP-2, Bromus inermis, Capsicum annuum L. var. annuum, Che MMP-3, MMP-9 and HLE. In a further embodiment, the nopodium quinoa, Chenopodium quinoa Subsp. Quinoa, plant extract is capable of inhibiting at least three of MMP-1, Chenopodium quinoa Willd. Chichorium endivia, Chicho MMP-2, MMP-3, MMP-9 and HLE. rium endivia Subsp. Endivia, Citrullus lanatus, Cornus 0064. The plant extracts may be selected from extracts sericea, Daucus carota, Daucus carota Subsp carota L., known in the art and subsequently tested for their ability to Dolichos lablab, Euphorbia amygdaloides, Foeniculum vul inhibit one or more of MMP-1, MMP-2, MMP-3, MMP-9 gare, Galinsoga quadriradiata, Gentiana lutea, Geranium and/or HLE, or they may be identified using the process sanguineum, Geraniumxcantabrigiense, Glycyrrhiza gla bra, Helianthus strumosus, Hypomyces lactifluorum, Juni described herein. In one embodiment of the present inven perus communis L., Lentinus edodes, Lotus corniculatus, tion, the plant extracts are derived from one of the plants Manihot esculenta, Matricaria recutita, Melilotus albus, listed in Tables 1 to 5. In another embodiment, the plant Melilotus alba Medik, Melissa officinalis, Oenothera bien extracts are derived from one of the plants listed in Table 6. nis, Pastinaca sativa L., Phaseolus vulgaris, Physalis phila 0065. In another embodiment, the plant extracts are delphica, Pimpinella anisum, Pisum sativum, Potentilla derived from a: plant selected from the group of Aconitum anserina L., Potentilla fruticosa, Raphanus raphanistrum, napellus, Acorus calamus, Alchemilla mollis, Allium cepa, Rheumxhybridum, Rhus typhina L., Ribes Sylvestre, Rodg Allium sativum, Allium tuberosum, Ambrosia artemisiifolia, ersia spp., Rubus Occidentalis, Rubus thibetanus, Rumex Anethum graveolens, Anthemis tinctoria, Aronia melano crispus, Rumex scutatus, Setaria italica, Solanum melon carpa (MichX) Ell. Arctostaphylos uva-ursi, Aroniaxpruni gena L., Sorghum dochna bicolor gr technicum, Stellaria folia, Artemisia dracunculus, Avena sativa, Beta vulgaris, media, Tanacetum cinerariifolium, Taraxacum officinale, Beta vulgaris L. Subsp. Vulgaris, Borago officinalis, Bras Thymus fragantissimus, ThymusXcitriiodorus, Trifolium sica napus, Brassica oleracea, Brassica oleracea L. var. incarnatum, Triticosecale spp., Tropaeolum majus L., Tsuga italica Plenck, Brassica rapa, Bromus inermis, Capsicum Canadensis, Tsuga diversifolia, Vaccinium angustifolium, annuum L. var. annuum, Cerastium tomentosum, Chaero Vaccinium angustifolium Ait. Vitia sp., xTriticosecale spp., phyllum bulbosum, Chenopodium quinoa, Chenopodium Zea may’s L. and Zingiber officinale quinoa Subsp. Quinoa, Chenopodium quinoa Willd. Chi 0067. In another embodiment of the present invention, chorium endivia, Chichorium endivia subsp. Endivia, Cir the plant extract is derived from a plant selected from the cium arvense, Citrullus lanatus, Cornus Canadensis, Cornus group of Beta vulgaris L., Brassica oleracea L., Capsicum sericea, Cynara Cardunculus subsp. Cardunculus, Daucus annuum L. Chenopodium quinoa, Daucus carota L., Gera carota, Daucus carota Subsp carota L., Dolichos lablab, niumxcantabrigiense, Juniperus communis L., Melilotus Euphorbia amygdaloides, Fagopyrum tataricum, Foenicu alba, Pastinaca sativa L., Potentilla anserina L., Rhus lum vulgare, Frangula alnus, Galinsoga quadriradiata, typhina L., Solanum melongena L., Triticosecale spp., Tro Gentiana lutea, Geranium sanguineum, Geraniumxcant paeolum maius L., Vaccinium angustifolium, and Zea mays abrigiense, Glycyrrhiza glabra, Hamamelis virginiana, L. Helianthus strumosus, Heliotropium arborescens, Hordeum vulgare Subsp. Vulgare, Hypomyces lactifluorum, Juniperus 0068. In accordance with the present invention, the plant communis L., Lentinus edodes, Lotus corniculatus, Manihot extracts are solvent-based extracts obtained from the esculenta, Matricaria recutita, Melilotus albus, Melilotus selected plant by solvent extraction. The solvent can be an alba Medik, Melissa officinalis, Menthaxpiperita, aqueous solvent (such as water or a buffer), or it can be a Oenothera biennis, Pastinaca sativa L., Petroselinum liquid organic compound, or a combination of an aqueous crispum, Phaseolus vulgaris, Physalis philadelphica, Phy Solvent and a liquid organic compound. In one embodiment tolacca decandra, Phytolacca decandra syn. P. americana, of the invention, the plant extract is an aqueous, alcoholic or Pimpinella anisum, Pisum sativum, Potentilla anserina L., aqueous alcoholic extract. In another embodiment, the plant Potentilla fruticosa, Poterium sanguisorba, Pyrus commu extract is an aqueous, ethanolic, glycolic, aqueous-ethanolic nis, Raphanus raphanistrum, Rheumxhybridum, Rhus or aqueous-glycolic extract. In a further embodiment, the typhina L., Ribes nigrum L. Ribes Sylvestre, Rodgersia spp., glycol is butylene glycol. Rosmarinus officinalis, Rubus Occidentalis, Rubus thibeta nus, Rumex Crispus, Rumex scutatus, Ruta graveolens, 0069 Preparation of the Plant Extracts Salvia officinalis, Sambucus Canadensis L., Setaria italica, 0070 The plant extracts are obtained by solvent extrac Solanum melongena L., Sorghum dochna bicolor gr techni tion of plant material from a selected plant. The actual cum, Stellaria media, Tanacetum cinerariifolium, Tarax extraction process is not critical to the invention, but typi acum officinale, Teucrium chamaedrys, Thymus fragantissi cally employs as solvent an aqueous solvent (such as water initiS, ThymusXcitriodorus, Trifolium incarnatun, of a buffer), a liquid organic compound, or a combination Triticosecale spp., Tropaeolum maius L., Tsuga Canadensis, thereof. Exemplary liquid organic compounds that can be US 2007/01 22492 A1 May 31, 2007 used as solvents in the extraction process to prepare the plant 0075) A number of standard extraction techniques known extracts include, but are not limited to, primary alcohols in the art can be employed to prepare the plant extracts. In Such as methyl alcohol (methanol), ethyl alcohol (ethanol), general, the extraction process entails contacting Solid plant 1-propanol and 1-butanol; secondary alcohols such as 2-pro material with a solvent with adequate mixing and for a panol and 2-butanol; tertiary alcohols such as 2-methyl-2- period of time sufficient to ensure adequate exposure of the propanol; liquid polyhydric alcohols such as glycerine and solid plant material to the solvent such that inhibitory glycols; and other known organic solvents such as acetone, activity present in the plant material can be taken up by the tetrahydrofuran, acetonitrile, 1,4-dioxane, pyridine, dimeth solvent. ylsulfoxide, N,N-dimethyl formamide, acetic acid, diethyl ether, hexane, heptane, dichloromethane and ethyl acetate. 0076. The plant material employed in the extraction Suitable glycols include, for example, ethylene glycol, pro process can be the entire plant, or it can be one or more pylene glycol, diethylene glycol, dipropylene glycol and distinct tissues from a plant, for example, leaves, flowers, 1,3-butylene glycol. roots, seeds, pods, stems, fruits, seed coats, buds, or various combinations thereof. The plant material can be fresh, dried 0071. When the extraction process is carried out using a or frozen. The plant material may be used immediately after Solvent that comprises a mixture of an aqueous solvent and harvesting or it can be stored for a period of time prior to a liquid organic compound, the content of the liquid organic being Subjected to the extraction process. If the plant mate compound ranges from about 5% to about 95% by volume. rial is stored, it can be treated prior to storage, for example, In one embodiment, the content of the liquid organic com by drying, freezing, lyophilising, or some combination pound in the solvent ranges from about 10% to about 90% thereof. The storage time may be of various durations, for by volume. In another embodiment, the content of the liquid example, the storage period may be between a few days and organic compound in the solvent ranges from about 20% to a few years. Typically storage times range between less than about 90% by volume. In a further embodiment, the content one week to about one year in duration. of the liquid organic compound in the solvent ranges from about 20% to about 85% by volume. In another embodi 0077. If desired, the plant material can be derived from a ment, the content of the liquid organic compound in the plant that was subjected to a harvest stress treatment. A solvent ranges from about 20% to about 50% by volume. In stress treatment comprises contacting or treating the plant, an alternate embodiment, the content of the liquid organic or material from the plant, with one or more stressor with the compound in the solvent ranges from about 50% to about aim of inducing or eliciting increased production of one or more chemicals. The stressor can be a chemical compound 85% by volume. or a physical treatment. Examples of chemical stressors 0072 For dermatological applications wherein the plant include, but are not limited to, organic and inorganic acids extract will be formulated for topical use, a solvent that is including fatty acids, glycerides, phospholipids, glycolipids, compatible with mammalian skin can be selected. Examples organic solvents, amino acids, peptides, monosaccharides, of Such solvents include, but are not limited to, water, an oligosaccharides, polysaccharides, lipopolysaccharides, aqueous buffer, a combination of water/buffer and a lower phenolics, alkaloids, terpenes, terpenoids, antibiotics, deter alcohol or an anhydrous lower alcohol. In the context of the gents, polyamines, peroxides, ionophores, and the like. present invention, a lower alcohol refers to an alcohol Examples of physical stress treatments include, but are not having 1 to 4 carbon atoms, such as a primary, secondary, limited to, ultraviolet radiation, Sandblasting, low and high tertiary or liquid polyhydric alcohol. Accordingly, in one temperature stress, and osmotic stress induced by Salt or embodiment of the present invention, the solvent is selected Sugars. Nutritional stress is defined as depriving the plant of from water, a lower alcohol or a combination thereof. In essential nutrients (e.g. nitrogen, phosphorus or potassium) another embodiment, the lower alcohol is selected from the in order to induce or elicit increased production of one or group of methyl alcohol (methanol), ethyl alcohol (ethanol), more chemicals. The one or more stressor (i.e. chemical 1-propanol, 1-butanol, 2-propanol, 2-butanol, 2-methyl-1- compound(s), physical treatment(s), or combination thereof) propanol, 2-methyl-2-propanol, glycerine, ethylene glycol, may be applied continuously or intermittently to the plant propylene glycol, diethylene glycol, dipropylene glycol and material. Various stressors and procedures for stressing 1,3-butylene glycol. plants prior to extract preparation have been described previously (see International Patent Application WO 0073. When the extraction employs a combination of an 02/06992) and are suitable for use in the present invention. aqueous solvent and a lower alcohol as solvent, the lower alcohol content of the solvent typically ranges from about 0078. In one embodiment of the present invention, the 10% to about 95% by volume. In one embodiment of the plant extract is prepared from a plant that has been Subjected present invention, the lower alcohol content of the solvent to a stress treatment. In another embodiment, the extract is ranges from about 10% to about 90% by volume. prepared from a plant that has been Subjected to one or more chemical stressors. In a further embodiment, the extract is 0074. In another embodiment, the lower alcohol content prepared from a plant that has been Subjected to one or more of the solvent ranges from about 15% to about 90% by chemical stressors selected from the group of Y-linolenic volume. In a further embodiment, the lower alcohol content acid, Y-linolenic acid lower alkyl esters, arachidonic acid and of the solvent ranges from about 15% to about 50% by arachidonic acid lower alkyl esters. In a further embodiment, volume. In another embodiment, the lower alcohol content the extract is prepared from a plant that has been Subjected of the solvent ranges from about 50% to about 90% by to one or more physical stress. In yet another embodiment, Volume. In an alternate embodiment, the solvent comprises the extract is derived from an unstressed plant. a combination of an aqueous solvent and a lower alcohol, wherein the lower alcohol content is not less than 20% by 0079 If desired, the plant material can be treated prior to Volume. the extraction process in order to facilitate the extraction US 2007/01 22492 A1 May 31, 2007 process. Typically such treatment results in the plant mate MMP-9 and HLE, using a variety of techniques known in rial being fragmented by some means such that a greater the art including, but not limited to, those described herein. Surface area is presented to the solvent. For example, the In the context of the present invention, a plant extract that plant material can be crushed or sliced mechanically, using decreases the activity of an EP by at least 20% is considered a grinder or other device to fragment the plant parts into to be capable of inhibiting the activity of that protease. In Small pieces or particles, or the plant material can be frozen one embodiment of the present invention, a plant extract that in liquid nitrogen and then crushed or fragmented into inhibits the activity of one or more of MMP-1, MMP-2, Smaller pieces. MM-3, MMP-9 and HLE by at least 20% is considered to be an extract of the invention. In another embodiment, the plant 0080. The amount of the solvent used in the extraction extract inhibits the activity of one or more of MMP-1, can range from about 1x to about 100x (mass/mass) that of MMP-2, MMP-3, MMP-9 and HLE by at least 30%. In the solid plant material. In one embodiment of the present another embodiment, the plant extract inhibits the activity of invention, the amount of Solvent used in the extraction one or more of MMP-1, MMP-2, MMP-3, MMP-9 and HLE process ranges from about 1x to about 50x (mass/mass) that by at least 40%. In a further embodiment, the plant extract of the solid plant material. inhibits the activity of one or more of MMP-1, MMP-2, 0081. A variety of conditions can be employed for the MMP-3, MMP-9 and HLE by at least 45%. In another extraction process. Typically, the extraction procedures are embodiment, the plant extract inhibits the activity of one or conducted over a period of time between about 10 minutes more of MMP-1, MMP-2, MMP-3, MMP-9 and HLE by at and about 24 hours at a temperature between about 4°C. and least 50%. about 50° C. However, temperatures between about 4° C. and about 90° C., for example between about 4° C. and 0088. In order to determine whether the extracts inhibit a about 70° C. can be employed. Similarly, extraction time skin EP, the extract can be tested against an individual skin may be varied depending on other extraction conditions, for EP or against a panel comprising two or more of MMP-1, example the extraction time can range from several minutes MMP-2, MMP-3, MMP-9 and HLE. to several hours. 0089. As indicated above, a variety of methods and techniques for measuring the ability of the extracts to inhibit 0082) Adequate contact of the solvent with the plant the activity of a skin EP either qualitatively or quantitatively material can be encouraged by shaking the Suspension. are known in the art. For example, there are currently several Alternatively, an extraction device equipped with, for assays to measure the activity of MMPs and elastase (for a instance, a stirring machine, can be employed which may review of these methods, see Murphy and Crabbe. In Barrett improve the extraction efficiency. The extraction can be (ed.) Methods in Enzymology. Proteolytic Enzymes. Aspartic carried out at ordinary pressure, under pressure or at reduced Acid and Metallopeptidases, New York: Academic Press, pressure established by, for example, aspiration. Appropriate 1995, 248: 470), including the gelatineolytic assay (which is extraction conditions can readily be determined or selected based on the degradation of radio-labelled type I collagen), by one skilled in the art taking into consideration the the Zymography assay (which is based on the presence of production conditions such as production facilities and negatively-stained bands following electrophoresis through yields. Substrate-impregnated SDS polyacrylamide gels) and a 0083. Following the extraction process, the liquid frac microtitre plate assay developed by Pacmenet al., (Biochem. tion (the primary plant extract) can be separated from the Pharm. (1996) 52:105-111). solid (insoluble) matter. Separation of the liquid and solid fractions can be achieved by one or more standard separa 0090. Other methods include those that employ auto tion processes known to those skilled in the art, such as quenched fluorogenic Substrates. Many fluorogenic Sub various centrifugation or filtration processes. strates have been designed for quantification of the activity of MMPs and elastase, through fluorescent level variation 0084. If desired, the primary extract can be subjected to measuring (reviewed by Nagase and Fields (1996) Biopoly one or more additional steps to further purify the extract. For mers 40: 399-416). Another method of measuring EP activ example, the primary extract may be subjected to Solid ity makes use of the fluorescent activated substrate conver liquid extraction, liquid-liquid extraction, Solid-phase sion (FASC) assay described in Canadian Patent No. 2,189, extraction (SPE), membrane filtration, ultrafiltration, dialy 486 and in St-Pierre et al., (1996) Cytometry 25:374-380). sis, electrophoresis, solvent concentration, centrifugation, ultracentrifugation, liquid or gas phase chromatography 0091 Various formats known in the art may be employed (including size exclusion, affinity, etc.) with or without high in the assays. For example, the extract may be tested against pressure, lyophilisation, evaporation, precipitation with one or more EPs in a sequential fashion or it may be tested various “carriers” (including PVPP carbon, antibodies, and against a plurality, or array, of skin EPs simultaneously. The the like), the use of supercritical fluids (such as CO), or assays may be adapted to high throughput as is known in the various combinations thereof to provide a substantially pure art in order to facilitate simultaneous testing of an extract eXtract. against a plurality of skin EPs. 0092. The assays can be conducted using purified or 0085 Testing the Plant Extracts semi-purified EPs. Methods of isolating and purifying EPs 0086) Determination of Extracellular Protease Inhibitory are well known in the art. In addition, many EPs are Activity commercially available (for example, from Sigma-Aldrich, 0087. Following the extraction process, the plant extract St. Louis, Mo. and Calbiochem, San. Diego, Calif.). can be tested for its ability to inhibit one or more skin EPs 0093. Alternatively, the ability of the extracts to inhibit selected from the group of MMP-1, MMP-2, MMP-3, the activity of skin EPs can be evaluated using cultures of US 2007/01 22492 A1 May 31, 2007

cells that secrete one or more skin EPs. In this case a cell 0100. In order to assess the protease activity in skin culture is contacted with an appropriate amount of the biopsies, the samples are typically flash frozen, mechani extract. After an appropriate period of time, the cells are cally ground and/or homogenised. After centrifugation, the extracted, centrifuged and the proteolytic activity in the Supernatants are isolated and used to assess EP activity in Supernatant is measured. This method is useful in determin assays such as those outlined above. ing the ability of an extract to inhibit a set of EPs secreted by a particular cell line or combination of cell lines. For 0101 In vitro Cellular Activity in Skin Cells example, assays can be conducted with cell lines derived 0102) In order to be useful in dermatological applica from mammalian skin, such as keratinocytes or fibroblasts. tions, the selected plant extracts are capable of affecting one or more cellular activity of skin cells in a beneficial manner. 0094) Inhibition of EPActivity in Skin Models The ability of plant extracts to affect one or more cellular 0.095 As an extension of the cell culture assays described activities in skin cells can be assessed in vitro using one, or above, the extracts may be tested in an appropriate skin a combination, of standard techniques known in the art. model for their ability to inhibit one or more of MMP-1, Cellular activities in skin cells that can be assessed in vitro MMP-2, MMP-3, MMP-9 and HLE. For example, an in include, but are not limited to, the breakdown of a structural vitro human skin model can be employed to test the component of the ECM, such as collagen, fibronectin, fibril extract(s). Such models are typically constructed from lin and/or elastin; cell migration; collagen production; UV human fibroblasts and keratinocytes by first forming a gel induced extracellular protease activity and tractional forces comprising human dermal fibroblasts and collagen. Cell generated by fibroblasts; response to oxidative stresses, culture medium is added and the gel incubated for a suffi inhibition of release of IL-8 or other cytokines, response to cient number of days to allow for fibroblast proliferation, induced apoposis, wound healing. and for collagen and protease synthesis and secretion into 0.103 For example, the ability of the extracts of the the gel. Following this incubation period, donor-matched invention to attenuate the breakdown of one or more ECM human epidermal keratinocytes in a biological medium are component can be assessed in vitro using skin models such gently pipetted onto the gel and allowed to establish a as those described above. After incubation with a plant confluent layer on its surface. The test plant extract is added extract, the gels can be extracted and assayed for the loss of and after a suitable incubation period (for example, between one or more structural components of the ECM, such as 6 and 24 hours), the gels are extracted and centrifuged and elastin, collagen, fibronectin and/or fibrillin. Alternatively, the proteolytic activity in the Supernatant is assayed. the gels can be assayed for the presence of fragments of 0096. Immune cells can also be added to the above skin elastin, collagen, fibronectin and/or fibrillin using standard model in order to provide a source of elastase enzymes. techniques as an indication of the breakdown of these Other examples of skin models are provided in the art, for components. example, see U.S. Pat. No. 6,079,415 and references therein. 0.104 Elastin, for example, can be quantitated biochemi 0097 In vivo Testing of EPe Inhibition cally as desmosine or visualized histologically (Starcher B and Conrad M: Ciba Found Symp. (1995) 192:338-46). 0098. Alternatively, the ability of the extracts to inhibit Alternatively, confocal microscopy can be used in visualize skin EP activity may be assessed in vivo using various the dermal microfibrillar network (Watson RE et al: J Invest standard techniques. For example, the ability of the extracts Dermatol. (1999) 112(5):782-7). Intact elastin and elastin to inhibit protease activity can be determined in animal fragments can also be measured by immunoblotting models or human Volunteers. An example of a Suitable (Sakuraoka Ket al: J Dermatol Sci (1996) 12(3):232-237). animal model would be a skh-1 mouse or nude mouse or rat that is treated with an extract of the invention and then 0105 Biochemical and/or immunochemistry methods exposed to UV radiation (see, Nishimori et al. (2001) J. can be used to assess changes in the amount of collagen in Invest. Dernatol. 117:1458-1463). UV radiation is known to the gels. Ultrastructural methods can also be used to assess increase the level of activity of certain MMPs (see, for changes in the amount of collagen in the gels (Fligiel S E et example, U.S. Pat. No. 6,130,254). Skin biopsies are taken al:J Invest Dermatol. (2003) 120(5):842-8). Type I collagen, from the animal and the amount of EP activity in the the most abundant extracellular matrix protein deposited in biopsied sample can be measured using standard techniques cutaneous involvement, can be measured using the method as an indication of the inhibitory activity of the test extract. described by Allanore Yetal (J Rheumatol. (2003)30(1):68 73). 0099 Human trials may also be used to evaluate the ability of an extract to inhibit EP activity in the skin. For 0106 Quantitative reverse transcriptase-polymerase example, skin biopsies can be taken from adult Volunteers chain reaction analysis can be used to determine the pres exposed to UV radiation and treated prior to or after UV ence of dermal elastosis, diminished fibrillin and type VII exposure with an extract. The biopsy samples can be collagen expression (Bosset S et al: Br J Dermatol. (2003) assessed for EP activity and compared to an appropriate 148(4):770-8). control (for example, skin biopsies from individuals treated 0.107 Some of the more complex skin models allow for with a control compound or untreated individuals). Alterna more Sophisticated testing procedures such as those tively, as an age-related increase in the relative activities of described by Roguet R (Skin Pharmacol Appl Skin Physiol. MMP-1, MMP-2 and MMP-9 has been demonstrated (see, for example, U.S. Patent Application No. 20010053347), (2002) 15 Suppl 1:1-3), which can also be employed in elderly individuals (for example, those over 80 years of age) testing the plant extracts. could be used as volunteers for the trials without the 0108. In general, the ability of an extract to inhibit requirement for UV exposure. migration of cells can be assessed in vitro using standard cell US 2007/01 22492 A1 May 31, 2007 migration assays. Typically, Such assays are conducted in 0114. As is known in the art, MMPs may act to extend multi-well plates, the wells of the plate being separated by anchoring offibroblasts on the extracellular matrix, resulting a Suitable membrane into top and bottom sections. The in greater fibroblast tractional forces. Accordingly, the effect membrane is coated with an appropriate compound, the of the plant extracts on the tractional forces generated by selection of which is dependent on the type of cell being fibroblasts can be assayed. This assay employs a model assessed and can be readily determined by one skilled in the comprising fibroblasts embedded in a collagen matrix to art. Examples include collagen, gelatinee or Matrigel for create a derm-like environment. Such a model can be endothelial cells. An appropriate chemo-attractant, such as prepared by adding fibroblasts to a solution of collagen I in EGM-2, IL-8, C.FGF, BFGF and the like, is added to the medium and then allowing the collagen to polymerize to bottom chamber as a chemo-attractant. An aliquot of the test form a gel. After an appropriate incubation period, the cells together with the extract are added to the upper derm-like gel is treated with an extract and the amount of chamber. Typically various dilutions of the extract are tested. contraction measured over a period of time, for example, After a suitable incubation time, the membrane is rinsed, several days. The amount of contraction can be assessed for fixed and stained. The cells on the upper side of the example, by digitally photographing the gel at various time membrane are wiped off, and then randomly selected fields points and calculating the gel area using appropriate soft on the bottom side are counted. ware. The amount of contraction can be compared to 0109 Inhibition of cell migration can also be assessed untreated control gels and/or gels treated with a compound using the cord formation assay. In this assay endothelial cells known to affect fibroblast tractional forces. with or without plant extract are plated onto Matrigel and 0115 Additional Testing incubated under appropriate conditions. After a suitable period of time (for example, between 18 and 24 hours), 0.116) The plant extracts may undergo additional testing if migration of cells is assessed by visual inspection to deter desired. For example, the ability of the plant extracts to mine whether the cells have formed into cords. affect one or more cellular activity of skin cells can be assessed in vivo and/or the plant extracts may be submitted 0110 Various cell lines can be used in cell migration to testing on human Volunteers to assess their ability to exert assays. Examples of suitable endothelial cell lines include, the desired dermatological effect(s). The plant extracts may but are not limited to, human umbilical vein endothelial cells also undergo one or more safety, stability and/or bioavail (HUVECs), bovine aortic endothelial cells (BAECs), human ability test prior to testing on human Volunteers. coronary artery endothelial cells (HCAECs), bovine adrenal gland capillary endothelial cells (BCE) and vascular smooth 0.117) 1. In vivo Testing muscle cells. HUVECs can be isolated from umbilical cords 0118. The ability of the extracts of the invention to affect using standard methods (see, for example, Jaffe et al. (1973) one or more cellular activity of skin cells can be assessed in J. Clin. Invest. 52: 2745), or they can be obtained from the Vivo using various standard techniques. For example, using ATCC or various commercial sources, as can other Suitable appropriate animal models and/or human Volunteers. endothelial cell lines. 0111. The effect of the plant extracts on collagen I pro 0119) Degeneration of the ECM, in particular due to the duction in the skin cells can be assessed, for example, using breakdown of collagen and/or elastin, can be assessed in immunochemical methods. One exemplary method involves skin biopsies, for example, by histological examination of measuring the release of the procollagen type I C-peptide skin tissue after treatment with the extract. Methods (PIP) in skin cells treated with the extract and comparing this described above for the determination of the breakdown of to the amount of PEP released by untreated controls and/or one or more structural components of the ECM can also be controls treated with a compound known to affect collagen used on the biopsied samples. Histology can also be used to production. ELISA kits suitable for assaying PIP are com determine abnormal cell migration. mercially available (for example from Takara Mirus Bio, 0120 Skin changes, such as wrinkling and/or sagging, Madison, Wis.). As PIP is cleaved off the procollagen reddening, formation of lesions, abnormal pigmentation and molecule during formation of the collagen triple helix, the the like, can be assessed by visual examination. For amount of this peptide released by the skin cells is stoichio example, the effect of the plant extraction the skin can be metrically proportional to the amount of collagen synthe evaluated by formulating the extract such that it is suitable sized. for external application to the skin and Susequently sensory 0112 UV-induced extracellular protease activity can be tests can be conducted on the formulation using by a panel assessed by irradiating cultures of skin cells with UVA light of human Volunteers. A sensory test typically involves and then treating the irradiated cells with the extract. Alter application of the formulation to the skin of the panelists on natively, the extract can be added to the cells prior to a regular basis. Such as once or twice a day, over a period of irradiation to assess the prophylactic effect of the extract. several weeks. The effect of the formulation on the skin can After a suitable period of incubation in an appropriate be evaluated by inspecting the skin of the panelists and medium, Supernatants can be removed from the cells and assessing visually the skin characteristic or characteristics assayed for proteolytic activity as described above. Results being investigated, for example, the tenseness and gloss of can be compared to untreated cells and/or cells treated with the skin, a decrease of any wrinkles, sags, reddening, lesions a compound known to affect UV-induced protease activity. and/or abnormal pigmentation. 0113 Skin cells suitable for use in the above assays 0121 Erythema in skin samples can be determined, for include human dermal fibroblasts, keratinocytes, melano example, using commercially available chromameter. The cytes, Langerhans cells, cells of the hair follicle and cells of ability of the plant extracts to reduce inflammation in the the immune system which produce proteases, including skin can also be assessed in human Volunteers using stan leukocytes, macrophages and lymphocytes. dard techniques, including visual inspection. US 2007/01 22492 A1 May 31, 2007

0122) The ability of the plant extract to inhibit endothelial be a synthetic membrane, for example polysulphone, cellu cell migration can also be assessed in vivo, using standard lose acetate or nitrate, or polytetrafluoroethylene, or it can be techniques such as the CAM assay (see Brooks et al., in a skin Sample, Such as a sample taken from a cadaver. Methods in Molecular Biology, Vol. 129, pp. 257-269 (2000), ed. A. R. Howlett, Humana Press Inc., Totowa, N.J.; 0127. Other tests are known in the art (for example, see Ausprunk et al., (1975) Am. J. Pathol., 79:597-618; Osson U.S. Pharmacopoeia XXII (1990)) and are suitable for ski et al., (1980) Cancer Res., 40:2300-2309), the Matrigel testing the stability and/or safety of the extracts. plug assay (see, for example, Passaniti, et al., (1992) Lab. 0128. As will be readily apparent to one skilled in the art, Invest. 67:519-528) or the corneal micropocket assay (see a selected extract may need to meet certain criteria in order D'Amato, et al., (1994) Proc. Natl. Acad. Sci. USA, to meet regulatory requirements for human use. Conducting 91:4082-4085; Kochet al., (1991) Agents Actions, 34:350-7: tests such as those described above, therefore, allows the Kenyon, et al., (1996) Invest. Ophthalmol. Vis. Sci. 37:1625 Suitability of an extract for human use to be assessed. 1632). 0.129 Isolation of Active Ingredients 0123 The CAM assay measures neovascularization of whole tissue, wherein chick embryo blood vessels grow into 0.130. The present invention also provides for active the CAM or into the tissue transplanted on the CAM, and is ingredients isolated from the plant extracts of the invention. a well-recognised assay model for in vivo angiogenesis. The In the context of the present invention an “active ingredient’ Matrigel plug assay involves introducing an extract into cold is a compound or molecule that is capable of inhibiting one liquid Matrigel which, after Subcutaneous injection into a or more skin EPs selected from the group of MMP-1, Suitable animal model, Solidifies and permits penetration by MMP-2, MMP-3, MMP-9 and HLE. The active ingredient host cells and the formation of new blood vessels. After a may be proteinaceous or non-proteinaceous. Isolated active suitable period of time, the animal is sacrificed, the Matrigel ingredients can be tested for their ability to inhibit one or plug is recovered and angiogenesis is assessed in the Matri more of MMP-1, MMP-2, MMP-3, MMP-9 and HLE using gel plug by measuring haemoglobin or by scoring selected the procedures described above. regions of histological sections for vascular density. Modi fications of this assay have also been described (see, for 0131 There are a number of techniques well known in example, Akhtar et al., (2002) Angiogenesis 5:75-80: Kragh the art for isolating active components from mixtures that et al., (2003) Int J Oncol. 22:305-11). The corneal micro may be employed to isolate the active ingredients from a pocket assay involves preparing pellets from a sterile hydron plant extract of the invention. These techniques include, but polymer containing a suitable amount of the extract. The are not limited to, Solid-liquid extraction, liquid-liquid pellets are Surgically implanted into corneal stromal micro extraction, solid-phase extraction (SPE), membrane filtra pockets created at an appropriate distance medial to the tion, ultrafiltration, dialysis, electrophoresis, solvent concen lateral corneal limbus of a test animal. Angiogenesis can be tration, centrifugation, ultracentrifugation, liquid or gas quantitated at various times after pellet implantation through phase chromatography (including size exclusion, affinity, the use of stereomicroscopy. Typically, the length of neoves and the like) with or without high pressure, lyophilisation, sels generated from the limbal vessel ring toward the centre evaporation, precipitation with various "carriers' (including of the cornea and the width of the neovessels are measured. PVPP carbon, antibodies, and the like), or various combi nations thereof. One skilled in the art, would appreciate how 0.124 2. Other Tests to use such options, in a sequential fashion, in order to enrich each successive fraction in the activity of interest by fol 0125. In addition to the above tests, the plant extracts of lowing its activity throughout the isolation procedure. the invention may be submitted to other standard tests to evaluate safety, cytotoxicity, stability, bioavailability and the 0132) Solid-liquid extraction means include the use of like. Exemplary tests to determine the cytotoxicity of the SOXhlet extractors, Vortex shakers, ultrasounds and other extracts and their potential to induce cytokine release are means to enhance extraction, as well as recovery by filtra described herein (see Examples X and XII). tion, centrifugation and related methods as described in the literature (see, for example, R. J. P. Cannell, Natural Prod 0126 The ability of an extract to penetrate the skin can be ucts Isolation, Humana Press, 1998). Examples of solvents assessed, for example, by in vitro release tests (see, for that may be used include, but are not limited to, hydrocarbon example, the U.S. Center for Drug Evaluation and Research Solvents, chlorinated Solvents, organic esters, organic ethers, guidance document entitled "Guidance for Industry. Non alcohols, water, and mixtures thereof. The use of supercriti sterile Semisolid Dosage Forms. Scale-up and postapproval cal solvents is also contemplated and includes the use of changes: in vitro release testing and in vivo bioequivalence modifiers such as those described in V. H. Bright (Super documentation'). Typically, Such testing is conducted using critical Fluid Technology, ACS Symp. Ser. Vol. 488, ch. 22, an open chamber diffusion cell, such as a Franz cell, fitted 1999). with an appropriate membrane. The test extract is placed on the upper side of the membrane and kept occluded to prevent 0.133 Liquid-liquid extraction means include the use of Solvent evaporation and compositional changes. A receptor various mixtures of solvents known in the art, including fluid. Such as aqueous buffer or hydro-alcoholic medium, is Solvents under Supercritical conditions. Typical Solvents placed on the other side of the membrane in a receptor cell. include, but are not limited to, hydrocarbon solvents, chlo Diffusion of the active component across the membrane is rinated Solvents, organic esters, organic ethers, alcohols, monitored by assay of sequentially collected Samples of the water, various aqueous Solutions, and mixtures thereof. The receptor fluid. For the extracts of the invention, the assay liquid-liquid extraction can be effected manually, or it can be could comprise, for example, testing the ability of the semi-automated or completely automated, and the solvent collected sample to inhibit EP activity. The membrane can can be removed or concentrated by standard techniques in US 2007/01 22492 A1 May 31, 2007 the art (see, for example, S. Ahuja, Handbook of Biosepa 0.139. In the case of countercurrent chromatography rations, Academic Press, 2000). (CCC), the present invention includes the use of manual, 0134 Solid-phase extraction (SPE) techniques include semi-automated, and automated systems, and the use of the use of cartridges, columns or other devices known in the various solvents and solvent combinations necessary to art. The sorbents that may be used with such techniques effect fractionation and/or purification of active ingredients include, but are not limited to, silica gel (normal phase), (see, for example, W. D. Conway, R. J. Petroski, Modern reverse-phase silica gel (modified silica gel), ion-exchange Countercurrent Chromatography, ACS Symp. Ser. Vol. 593, resins, and fluorisil. The invention also includes the use of 1995). Solvent removal and/or concentration can be effected Scavenger resins or other trapping reagents attached to Solid by various means known in the art including, but not limited Supports derived from organic or inorganic macromolecular to, reduced pressure evaporation, evaporation, reduced pres materials to remove selectively active ingredients or other Sure distillation, distillation, and lyophilization. constituents from the extracts. 0140. The present invention includes the isolation of 0135 Membrane, reverse osmosis and ultrafiltration active ingredients by expanded bed chromatography, mov means include the use of various types of membranes known ing and simulated moving bed chromatography, and other in the art, as well as the use of pressure, vacuum, centrifugal related methods known in the art (see, for example, G. Sofer, force, and/or other means that can be utilised in membrane L. Hagel, Handbook of Process Chromatography, Academic and ultrafiltration processes (see, for example, S. Ahuja, Press, 1997 and S. Ahuja, Handbook of Bioseparations, Handbook of Bioseparations, Academic Press, 2000). Academic Press, 2000). 0136 Dialysis means include membranes having a 0.141. Selective precipitation means includes the use of molecular weight cut-off varying from less than about 0.5 various solvents and solvent combinations, the use of tem KDa to greater than about 50 KDa. The invention also perature changes, the addition of precipitant and/or modifi covers the recovery of active ingredients from either the ers, and/or modification of the pH by addition of base or acid dialysate or the retentate by various means known in the art to effect a selective precipitation of active ingredients or including, but not limited to, evaporation, reduced pressure other constituents. evaporation, distillation, Vacuum distillation, and lyophiliza tion. 0142. The invention also includes the isolation of active ingredients by steam distillation, hydrodistillation, or other 0137 Chromatographic means include various means of related methods of distillation known in the art (see, for carrying out chromatography known by those skilled in the example, L. M. Harwood, C. J. Moody, Experimental art and described in the literature (see, for example, G. Sofer, Organic Chemistry, Blackwell Scientific Publications, UK, L. Hagel, Handbook of Process Chromatography, Academic Press, 1997). Examples include, but are not limited to, 1989). regular column chromatography, flash chromatography, high 0.143 Dermatological Formulations performance liquid chromatography (HPLC), medium pres sure liquid chromatography (MPLC), supercritical fluid 0144. The present invention further provides for formu chromatography (SFC), countercurrent chromatography lations Suitable for dermatological applications comprising (CCC), moving bed chromatography, simulated moving bed one or more extract of the invention, one or more active chromatography, expanded bed chromatography, and planar ingredient, or a combination thereof. The formulations can chromatography. With each chromatographic method, optionally comprise other therapeutic or cosmetic agents. examples of Sorbents that may be used include, but are not 0145 The formulations are prepared by standard tech limited to, silica gel, alumina, fluorisil, cellulose and modi niques such that they have acceptable toxicity and stability. fied cellulose, various modified silica gels, ion-exchange In addition, if the formulation is to be administered by a resins, size exclusion gels and other sorbents known in the route other than topical (e.g. systemic routes, such as oral, or art (see, for example, T. Hanai, HPLC: A Practical Guide, via intraperitoneal, intravenous, Subcutaneous and intramus RSC Press, UK 1999). The present invention also includes cular injection), then the extract and/or active ingredient the use of two or more solvent gradients to effect the must demonstrate acceptable hepatotoxicity and must be fractionation, partial purification, and/or purification of the Sufficiently resistant to degradation to allow the site of action active ingredients by chromatographic methods. Examples to be reached. of solvents that may be utilised include, but are not limited to, hexanes, heptane, pentane, petroleum ethers, cyclohex 0146 Testing for the above parameters and preparation of ane, heptane, diethyl ether, methanol, ethanol, isopropanol, appropriate formulations can be readily achieved by one propanol, butanol, isobutanol, tert-butanol, water, dichlo skilled in the art. Criteria which must be considered in the romethane, dichloroethane, ethyl acetate, tetrahydrofuran, preparation of a formulation include, but are not limited to, dioxane, tert-butyl methyl ether, acetone, and 2-butanone. the physicochemical and biochemical characteristics (bio When water or an aqueous phase is used, it may contain availability, toxicity, stability, etc.) of the extracts and/or varying amounts of inorganic or organic salts, and/or the pH active ingredients which make up the formulation. In par may be adjusted to different values with an acid or a base ticular, the formulation is prepared so as to preserve, as Such that fractionation and/or purification is enhanced. much as possible, the maximum inhibitory activity of the active components upon administration, without being 0138. In the case of planar chromatography, the present harmful to the animal. invention includes the use of various forms of this type of chromatography including, but not limited to, one- and two 0147 The formulations are prepared by mixing the dimension thin-layer chromatography (1D- and 2D-TLC), extract(s) and/or active ingredients together with a physi high performance thin-layer chromatography (HPTLC), and ologically acceptable carrier. Excipients, binders, diluents, centrifugal thin-layer chromatography (centrifugal TLC). and the like can also be included in the formulation. The US 2007/01 22492 A1 May 31, 2007

extract(s) and/or active ingredients can be formulated inde tablets may be uncoated or they may be coated by known pendently if desired and the respective formulations subse techniques to delay disintegration and absorption in the quently combined using a diluent or the like and adminis gastrointestinal tract and thereby provide a Sustained action tered, or can be administered independently of each other, over a longer period. For example, a time delay material either concurrently or at staggered times to the Subject. Such as glyceryl monostearate or glyceryl distearate may be 0148. The formulations according to the invention may employed. be in solid, semisolid or liquid form and may be adapted for 0151. Formulations for oral use may also be presented as oral (capsules, tablets, phials, troches, and the like), hard gelatinee capsules wherein the extract(s) and/or active parenteral, rectal, inhalation, or topical administration, and ingredients are mixed with an inert solid diluent, for may be in unit dosage form. The formulation may be adapted example, calcium carbonate, calcium phosphate or kaolin, or for slow release in vivo as known in the art. The formula as Soft gelatinee capsules wherein the extract(s) and/or tions of the invention may be used in conventional form active ingredients are mixed with water or an oil medium, including, but not limited to, Solutions, syrups, troches, for example peanut oil, liquid paraffin or olive oil. lozenges, aqueous or oily Suspensions, dispersible powders 0152 Aqueous suspensions contain extract(s) and/or or granules, emulsions, hard or soft capsules, elixirs, active ingredients in admixture with excipients suitable for injectables, tablets, capsules, Suppositories, hydrophobic the manufacture of aqueous Suspensions. Such excipients and hydrophilic creams and lotions. The term parenteral as are suspending agents, for example, Sodium carboxymeth used herein includes Subcutaneous injections, intravenous, ylcellulose, methyl cellulose, hydropropylmethylcellulose, intrathecal, intramuscular, intrasternal injection or infusion Sodium alginate, polyvinylpyrrolidone, gum tragacanth and techniques. gum acacia: dispersing or wetting agents may be a naturally 0149 Various physiologically acceptable carriers known occurring phosphatide, for example, lecithin, or condensa in the art can be used in the dermatological formulations of tion products of an alkylene oxide with fatty acids, for the invention. Examples of suitable carriers include, but are example polyoxyethyene Stearate, or condensation products not limited to, hydroxypropyl cellulose, starch (corn, potato, of ethylene oxide with long chain aliphatic alcohols, for rice, wheat), pregelatinized starch, gelatine, Sucrose, acacia, example hepta-decaethyleneoxycetanol, or condensation alginic acid, Sodium alginate, guar gum, ethyl cellulose, products of ethylene oxide with partial esters derived from carboxymethylcellulose sodium, carboxymethylcellulose fatty acids and a hexitol Such as polyoxyethylene Sorbitol calcium, polyvinylpyrrolidone, methylcellulose, hydrox monooleate, or condensation products of ethylene oxide ypropyl methylcellulose, microcrystalline cellulose, poly with partial esters derived from fatty acids and hexitol ethylene glycol, powdered cellulose, glucose, croScarmel anhydrides, for example polyethylene Sorbitan monooleate. lose Sodium, crospovidone, polacrilin potassium, Sodium The aqueous Suspensions may also contain one or more starch glycolate, tragacanth, calcium carbonate, dibasic cal preservatives, for example ethyl, or n-propyl p-hydroxy cium phosphate, tribasic calcium phosphate, kaolin, manni benzoate, one or more colouring agents, one or more fla tol, talc, cellulose acetate phthalate, polyethylene phthalate, vouring agents or one or more Sweetening agents, such as shellac, titanium dioxide, carnauba wax, microcrystalline Sucrose or saccharin. wax, calcium Stearate, magnesium Stearate, castor oil, min 0153. Oily suspensions may be formulated by suspending eral oil, light mineral oil, glycerine, Sorbitol, mannitol, the extract(s) and/or active ingredients in a vegetable oil, for Stearic acid, Sodium lauryl Sulfate, hydrogenated vegetable example, arachis oil, olive oil, Sesame oil or coconut oil, or oil (for example. peanut, cottonseed, Sunflower, Sesame, in a mineral oil such as liquid paraffin. The oily Suspensions olive, corn, soybean), Zinc Stearate, ethyl oleate, ethyl lau may contain a thickening agent, for example beeswax, hard rate, agar, calcium silicate, magnesium silicate, silicon diox paraffin or cetyl alcohol. Sweetening agents such as those set ide, colloidal silicon dioxide, calcium chloride, calcium Sulfate, silica gel, castor oil, diethyl phthalate, glyercin, forth above, and flavouring agents may be added to provide mono- and di-acetylated monoglycerides, propylene glycol, palatable oral preparations. These formulations may be triacetin, alamic acid, aluminum monostearate, bentonite, preserved by the addition of an anti-oxidant such as ascorbic bentonite magma, carbomer 934, carboxymethylcellulose acid. Sodium 12, carrageenan, hydroxyethyl cellulose, magne 0154 Dispersible powders and granules suitable for sium aluminum silicate, pectin, polyvinyl alcohol, povidine, preparation of an aqueous Suspension by the addition of Sodium alginate, tragacanth, Xanthan gum, and silicones. water provide the extract(s) and/or active ingredients in admixture with a dispersing or wetting agent, Suspending 0150. Formulations intended for oral use may be pre agent and one or more preservatives. Suitable dispersing or pared according to methods known in the art and may wetting agents and Suspending agents are exemplified by contain one or more agents such as Sweetening agents, those described above. Additional excipients, for example, flavouring agents, colouring agents and preserving agents in Sweetening, flavouring and colouring agents, may also be order to provide elegant and palatable preparations. Tablets present. contain the extract(s) and/or active ingredients in admixture with non-toxic physiologically acceptable excipients that are 0.155 Formulations of the invention may also be in the suitable for the manufacture of tablets. These excipients may form of oil-in-water emulsions. The oil phase may be a be, for example, inert diluents, such as calcium carbonate, vegetable oil, for example, olive oil or arachis oil, or a Sodium carbonate, lactose, calcium phosphate or sodium mineral oil, for example liquid paraffin or mixtures of these. phosphate: granulating and disintegrating agents for Suitable emulsifying agents may be naturally-occurring example, corn starch, or alginic acid: binding agents, for gums, for example, gum acacia or gum tragacanth, naturally example starch, gelatinee or acacia, and lubricating agents, occurring phosphatides, for example Soybean, lecithin, and for example magnesium Stearate, Stearic acid or talc. The esters or partial esters derived from fatty acids and hexitol, US 2007/01 22492 A1 May 31, 2007 anhydrides, for example Sorbitan monoleate, and condensa droxybutanoic acid; phenyl 2-hydroxyacetic acid; phenyl tion products of the said partial esters with ethylene oxide, 2-methyl 2-hydroxyacetic acid; 3-phenyl 2-hydroxyacetic for example polyoxyethylene sorbitan monoleate. The emul acid; 2,3-dihydroxypropanoic acid; 2,3,4-trihydroxybu sions may also contain Sweetening and flavouring agents. tanoic acid; 2.3.4.5,6-pentahydroxyhexanoic acid; 2-hy droxydodecanoic acid; 2.3.4.5-tetrahydroxypentanoic acid; 0156 Syrups and elixirs may be formulated with Sweet 2,3,4,5,6,7-hexahydroxyheptanoic acid; diphenyl 2-hy ening agents, for example, glycerol, propylene glycol, Sor droxyacetic acid, 4-hydroxymandelic acid, 4-chloroman bitol or Sucrose. Such formulations may also contain a delic acid; 3-hydroxybutanoic acid, 4-hydroxybutanoic acid; demulcent, a preservative and flavouring and colouring 2-bydroxyhexanoic acid; 5-hydroxydodecanoic acid; 12-hy agents. The formulations can be in the form of a sterile droxydodecanoic acid; 10-hydroxy decanoic acid; 16-hy injectable aqueous or oleaginous Suspension. This suspen droxyhexadecanoic acid; 2-hydroxy-3-methylbutanoic acid; sion may be formulated according to methods known in the 2-hydroxy-4-methylpentanoic acid; 3-hydroxy-4-methoxy art using Suitable dispersing or wetting agents and Suspend mandelic acid, 4-hydroxy-3-methoxymandelic acid; 2-hy ing agents such as those mentioned above. The sterile droxy-2-methylbutanoic acid; 3-(2-hydroxyphenyl) lactic injectable preparation may also be sterile injectable solution acid; 3-(4-hydroxyphenyl) lactic acid; hexahydromandelic or Suspension in a non-toxic parentally acceptable diluent or acid; 3-hydroxy-3-methylpentanoic acid, 4-hydroxyde Solvent, for example as a solution in 1,3-butanediol. Among canoic acid-5-hydroxy decanoic acid; aleuritic acid; 2-hy the acceptable vehicles and solvents that may be employed droxypropanedioic acid; 2-hydroxybutanedioic acid; tannic are water, Ringer's solution and isotonic sodium chloride acid; salicylic acid; erythraric acid; threaric acid, arabiraric Solution. In addition, sterile, fixed oils are conventionally acid; ribaric acid; Xylaric acid; lyXaric acid; glucaric acid; employed as a solvent or Suspending medium. For this galactaric acid; mannaric acid; gularic acid; allaric acid; purpose various bland fixed oils may be employed including altraric acid; idaric acid; talaric acid; 2-hydroxy-2-meth synthetic mono- or diglycerides. In addition, fatty acids Such ylbutanedioic acid; citric acid, isocitric acid, agaricic acid, as oleic acid find use in the preparation of injectables. quinic acid, glucoronic acid, glucoronolactone, galactoronic 0157. In one embodiment of the present invention, the acid, galactoronolactone, uronic acids, uronolactones, ascor dermatological formulations are for oral administration. bic acid, dihydroascorbic acid, dihydroxytartaric acid, tropic Such formulations can be presented as, for example, cap acid, ribonolactone, gluconolactone, galactonolactone, Sules, cachets, tablets, aerosol sprays, powders, granules, gulonolactone, mannonolactone, citramalic acid; pyruvic creams, pastes, gels, ointments, or as a solution or a sus acid, hydroxypyruvic acid, hydroxypyruvic acid phosphate pension in an aqueous liquid, a non-aqueous liquid, an and esters thereof methylpyruvate, ethyl pyruvate, propyl oil-in-water emulsion, or a water-in-oil liquid emulsion. pyruvate, isopropyl pyruvate; phenyl pyruvic acid and esters thereof methyl phenyl pyruvate, ethyl phenyl pyruvate, 0158. The formulations contemplated by the present propyl phenyl pyruvate; formyl formic acid and esters invention include so-called herbal and nutraceutical formu thereof: methyl formyl formate, ethyl formyl formate, pro lations. For nutraceutical formulations comprising Solid pyl formyl formate; benzoyl formic acid and esters thereof; parts of plant(s), the plant(s) must be an edible plant. The methyl benzoyl formate, ethyl benzoyl formate and propyl extract(s) and/or active ingredients or plant parts can be used benzoyl formate: 4-hydroxybenzoyl formic acid and esters in these herbal remedies and nutraceutical formulations as thereof 4-hydroxyphenyl pyruvic acid and esters thereof; Solutions, purified Solutions, or dry powders. and 2-hydroxyphenyl pyruvic acid and esters thereof. It 0159. In another embodiment of the present invention, should be understood that one or more derivatives of the the dermatological formulations are for topical administra above acidic component, Such as esters or lactones or tion. Such formulations may be presented as, for example, pharmaceutically acceptable salts thereof, may also be used. aerosol sprays, powders, Sticks, granules, creams, liquid 0162 Examples of moisturizing agents that are hydro creams, pastes, gels, lotions, syrups, ointments, on Sponges phobic agents include, but are not limited to, ceramide, or cotton applicators, or as a solution or a suspension in an borage oil (linoleic acid), tocopherollinoleate, dimethicone, aqueous liquid, a non-aqueous liquid, an oil-in-water emul glycerinee, and mixtures thereof. Examples of moisturizing Sion, or a water-in-oil liquid emulsion. agents that are hydrophilic agents include, but are not 0160 Topical formulations intended for application to limited to, hyaluronic acid, sodium peroxylinecarbolic acid the skin, hair and/or nails can include one or more moistur (sodium PCA), wheat protein (Such as laurdimonium izing agents, i.e. an agent that facilitates hydration of the hydroxypropyl hydrolyzed wheat protein), hair keratin skin by inhibiting or preventing loss of water from the skin, amino acids, and mixtures thereof. Sodium chloride may that absorbs water from the atmosphere and hydrates the also be present, for example, when hair keratin amino acids skin, and/or that enhances the skin's ability to absorb water are included as a moisturizer. Other moisturizing agents that directly from the atmosphere. Moisturizing agents generally may be included in the formulations include primrose oil minimise or prevent the skin from drying and cracking. and flax seed oil. Moisturizers, when used, are typically present in an amount 0.163 The formulation may further optionally include from about 0.01 to 20 weight percent of the formulation. one or more of a cysteine component, magnesium compo 0161 Suitable moisturizing agents include acidic com nent, manganese component, selenium component, and cop ponents, hydrophobic agents, and hydrophilic agents, or per component. These components are known in the art to combinations thereof. Examples of moisturizing agents that impart beneficial effects to the skin, hair and/or nails. are acidic components include, but are not limited to, 0164. For example a cysteine component may assist in 2-hydroxyacetic acid (glycolic acid), 2-hydroxypropanoic thickening the dermis and Supplementing collagen and elas acid (lactic acid), 2-methyl 2-hydroxypropanoic acid; 2-hy tic tissue, which can lead to a reduction of wrinkles and other US 2007/01 22492 A1 May 31, 2007

skin conditions. An example of a suitable cysteine compo amount from about 0.1 to 50 weight percent. Vitamin A, nent is N-acetyl cysteine, or a pharmaceutically acceptable when included, is usually in the form of vitamin A palmitate. salt thereof, which can be included in the formulation in an Vitamin A can be included in topical formulations in an amount from about 1 to 10 weight percent. The copper amount from about 0.5 to 15 weight percent. Suitable component may contribute to the inhibition elastase activity. carotenoids include, for example, beta-carotene, canthaxan Various copper compounds, or pharmaceutically acceptable thin, Zeaxanthin, lycopen, lutein, crocetin, capsanthin, and salts thereof, are suitable for inclusion in the formulations. mixtures thereof. Carotenoids can be included in the formu For example, copper sebacate can be included in the for lation in an amount from about 0.1 to 5 weight percent. mulation in an amount from about 5 to 20 weight percent. 0169. Other skin benefit ingredients can also be option 0165. The optional manganese component can be one of ally included in the dermatological formulations of the a variety of manganese compounds, or pharmaceutically present invention. Examples of skin benefit ingredients acceptable salts thereof, for example, manganese ascorbate include, but are not limited to, Sunscreens and Sunblocks, or a manganese ascorbic acid complex, which can be essential fatty acids, retinoids, cell activators, blood-circu included in the formulation in an amount from about 0.5 to lation promoters, tanning agents, alpha or beta hydroxy 10 weight percent. Suitable magnesium compounds include acids, proteins, peptides and polysaccharides. magnesium ascorbate or magnesium ascorbic acid complex. 0170 Sunscreens and sunblocks include those materials The magnesium component can be included in the formu commonly employed to block ultraviolet light. Examples of lation in an amount from about 1 to 10 weight percent. Suitable Sunscreens and Sunblocks include, but are not Suitable selenium compounds include selenium complexed limited to, titanium dioxide, Zinc oxide, talc, red veterinary with an amino acid, for example, L-selenomethionine. The petrolatum, a cinnamate (such as octyl methoxycinnamate), Selenium component can be included in the formulation in a benzone (such as oxybenzone or 2-hydroxy-4-methoxy an amount from about 0.01 to 3 weight percent. benzophenone), a salicylate (such as homosalicylate or octyl 0166 The dermatological formulation can also include salicylate), a benzoic acid (such as para-aminobenzoic acid), one or more anti-inflammatory components which facilitate and a benzophenone (such as oxybenzophenone). Octyl inhibition or Suppression of inflammation on or in the skin methoxycinnamate and 2-hydroxy-4-methoxy benzophe or in adjacent bodily tissues and thereby helps to reduce none (also known as oxybenzone) are commercially avail redness and swelling of the skin. Examples of suitable able under the trademarks, Parsol MCXTM and BenZophe anti-inflammatory components include vitamin E and none-3TM, respectively. The exact amount of sunscreen derivatives thereof, Zinc, allantoin, glycyrrhetic acid, aZu employed in the formulations will vary depending upon the lene, mefenamic acid, phenylbutaZone, indometacin, ibu degree of protection desired from the Sun's UV radiation and profen, ketoprofen, e-aminocaproic acid, hydrocortisone, can be readily determined by one skilled in the art. panthenol and derivatives and salts thereof, Zinc oxide and 0171 Essential fatty acids (EFAs) are those fatty acids diclofenac sodium. The anti-inflammatory component, when which are essential for the plasma membrane formation of used, can be incorporated into the formulations of the all cells. In keratinocytes, EFA deficiency makes cells hyper present invention in an amount between about 0.001 to about proliferative. EFAS also enhance lipid biosynthesis in the 5 weight percent. epidermis and provide lipids used in barrier formation by the 0167 The formulation may also optionally comprise one epidermis. Examples of essential fatty acids that may be or more anti-oxidants to help neutralize free radicals and included in the formulations include linoleic acid, Y-olino minimise their effect on the skin. Anti-oxidants can be lenic acid, homo-y-linolenic acid, columbinic acid, eicosa enzymatic or non-enzymatic type. Examples include the (n-6.9,13)-trienoic acid, arachidonic acid, Y-linolenic acid, enzymatic anti-oxidants: Superoxide dismutase (SOD), cata timnodonic acid, hexaenoic acid and mixtures thereof. lase, and glutathione peroxidase, and the non-enzymatic anti-oxidants: Vitamin E (for example, tocopherol) and 0172 AZoles, such as climbazole, bifonazole, clotrimna derivatives thereof, Vitamin A (retinol), Vitamin C (ascorbic Zole, ketoconazole, miconazole, econazole, itraconazole, acid), carotenoids and derivatives thereof, echinacoside, fluconazole, terconazole, butoconazole, Sulconazole, liona caffeoyl derivatives, oligomeric proanthocyanidins or Zole and mixtures thereof, may also optionally be included proanthanols (such as those obtained from grape seed in the formulations. extract), green tea polyphenols, dibutyl hydroxytoluene, 0173 Cell activators include, for example, royal jelly, butyl hydroxyanisole, tannin and derivatives thereof such as photosensitizers, cholesterol and derivatives thereof, fetal gallic acid and ellagic acid, flavonoids such as flavone, calf blood extract, vitamin A, retinols and retinoids, citric catechin, quercetin and leucoanthocyanidin, quinones Such acid, lactic acid, tartaric acid, malic acid, glycolic acid, as ubiquinone and vitamin K, thiamines and salts thereof, Succinic acid, serine, glutamic acid, hydroxyproline, thean riboflavins such as riboflavin and riboflavin acetate, pyri ine, pyrrolidone carboxylic acid, yeast extract, Lactobacillus doxines such as pyridoxine hydrochloride and pyridoxine extract and Bifidobacterium bifidum extract. The cell acti dioctanoate, nicotinic acids Such as nicotinic acid anmide vator(s) can be incorporated into the formulations in an and benzyl nicotinate, bihirubin, mannitol, tryptophane, amount between about 0.001 and 5 weight percent. histidine and nordihydroguaiaretic acid. 0.174 Examples of blood circulation promoters are 0168 When vitamin C is included in the formulation, it cepharanthine, tocopherol and derivatives thereof, nicotinic can be in the form of ascorbyl palmitate, dipalmitate acid and derivatives thereof; nonanoic acid vanillylamide, L-ascorbate, sodium L-ascorbate-2-Sulphate, or an ascorbic capsaicine, Zingerone, cantharides tincture, ichthammol. salt, such as sodium, potassium, and calcium, or mixtures caffeine, tannic acid, C.-borneol, cyclandelate, cinnarizine, thereof. Vitamin C can be included in the formulations in an tolazoline, acetylcholine, Verapamil, Y-oryZanol, camphor, US 2007/01 22492 A1 May 31, 2007

hinokitiol, and enzymes such as lipases and papain. The to, dry skin; dandruff acne, keratosis; psoriasis: eczema: blood circulation promoter(s) can be incorporated into the pruritus; age spots; reduced skin moisture; spider veins; formulations in an amount between about 0.01 to 20 weight senile purpura; lentigines; melasmas; deepening of skin percent. lines; blotches; wrinkles; blemished skin; nodules; atrophy; 0175. The formulations of the present invention can rosacea; impetigo; elastotic changes characterized by leath further optionally comprise one or more thickener. A thick ery, course, rough, dry and yellowish skin; telangiecatic ener will usually be present in amounts from 0.1 to 20% by skin; hyperpigmented skin; hyperkeratotic skin; inflamma weight of the formulation. Exemplary thickeners are cross tory dermatoses; "bullous' diseases, such as epidermolysis linked polyacrylate materials available under the trademark bullosa; hair breakage; hair loss; weathering damage; thin CarbopolTM (B.F. Goodrich Company), xanthan gum, car ning of the hair; brittle nails; thinning nails; flaking nails and rageenan, gelatinee, karaya, pectin and locust bean gum. ridged nails. Under certain circumstances the thickening function may be 0183 Improving the health and/or appearance of the skin, accomplished by a moisturizer component of the formula hair and nails, includes, for example, eliminating or pre tion. For instance, silicone gums and esters such as glycerol venting the dark skin, melasma or ephelis generated or Stearate have dual functionality. formed due to a variety of causes such as exposure to 0176) Other adjunct minor components can also option ultraviolet rays, changes in the hormone balance and genetic ally be incorporated into the dermatological formulations, programs; lightening the dullness of the skin; improving the for example, colouring agents, opacifiers, perfumes and gloss and/or firmness of the skin; inhibiting or preventing preservatives (for example, imidazolidinyl urea, dimethyl the progress of the skin-ageing phenomenon; reducing imidazolidinone or diazolidinyl urea). Amounts of these minor blemishes; controlling dandruff; reducing redness or materials can range from 0.001% up to 20% by weight of the inflammation of the scalp, and the like. The dermatological formulation. formulations of the present invention can also be used to promote wound healing and/or decrease the risk of scarring. 0177. The dermatological formulations intended for topi cal application can be packaged in a Suitable container to Suit 0.184 In another embodiment, an effective amount one or the viscosity and intended use. For example, a lotion or fluid more plant extracts and/or active ingredients of the inven cream can be packaged in a bottle or a roll-ball applicator, tion, or a dermatological formulation comprising an effec a capsule, a propellant-driven aerosol device or a container tive amount of one or more plant extracts and/or active fitted with a pump suitable for finger operation. When the ingredients is administered to a mammal in order to attenu composition is a cream or paste, it can simply be stored in ate one or more undesirable structural changes in the skin, a non-deformable bottle or Squeeze container, Such as a tube Such as wrinkling and/or sagging of the skin, loss of skin or a lidded jar. elasticity, redness, inflammation, formation of lesions, thin ning of the epithelium, abnormal migration of cells within 0178 USE the skin (such as that which occurs during angiogenesis or 0179 The plant extracts of the invention and/or active inflammation), or various combinations thereof. ingredients derived from the extracts, and formulations comprising the extracts and/or active ingredients are suitable 0185. One embodiment of the present invention provides for use for the routine care of the skin, hair and/or nails, to for the use of an effective amount of one or more plant improve the health and/or appearance of the skin, hair and/or extracts and/or active ingredients of the invention, or a nails and in the treatment or prevention of a variety of dermatological formulation comprising an effective amount dermatological conditions. of one or more plant extracts and/or active ingredients as a skin care product. In the context of the present invention a 0180. In the context of the present invention, a dermato “skin care product” refers to a product intended for use in the logical condition is a condition present on one or more of the maintenance and optimization of skin health and preserva components of the integumentary system of a subject, Such tion, from the standpoint of appearance and function. In as the skin, hair or nails, that is caused by ageing or by another embodiment of the present invention, the skin care intrinsic or extrinsic factors. Intrinsic factors include, for product is an anti-ageing product. An anti-ageing product is example, the genetic make up of an individual as well as a product intended to use in attenuating or preventing skin pathological conditions that cause undesirable effects on the ageing due to intrinsic or extrinsic factors. Skin ageing skin, hair or nails. Extrinsic factors include, but are not phenomena include, for example, skin thinning, fine and limited to, Sunlight, radiation, air pollution, wind, cold, coarse skin wrinkling, sagging, loss of elasticity, and the dampness, heat, chemicals, Smoke, and Smoking. like. Accordingly, the present invention provides for the 0181. Thus, an effective amount of one or more plant administration of an effective amount one or more plant extracts and/or active ingredients of the invention, or a extracts and/or active ingredients of the invention, or a dermatological formulation comprising an effective amount dermatological formulation comprising an effective amount of one or more plant extracts and/or active ingredients can of one or more plant extracts and/or active ingredients to a be administered to a mammal as part of routine skin/hair/nail mammal in order to produce an anti-ageing effect. maintenance, in order to improve the health and/or appear 0186 By “effective amount it is meant an amount of the ance of the skin, hair and/or nails or in order to treat or plant extract or active ingredient that provides a beneficial prevent a dermatological condition. In one embodiment of effect in the treatment of a dermatological condition or a the present invention, the plant extracts, active ingredients or desired skin improvement effect. It should be understood by formulations are administered topically to a mammal. one of ordinary skill in the art that this amount will vary 0182 Examples of dermatological conditions contem depending on the application and on the individual subject plated by the present invention include, but are not limited and will be readily determinable by one of skill in the art. US 2007/01 22492 A1 May 31, 2007

0187. Appropriate doses of a formulation comprising the 0.195 The extracts to be screened are prepared from plant plant extract(s) and/or active ingredient will also vary material derived from a plant or plants of interest, i.e. according to the age, body weight, and response of the “potential plants. Potential plants include all species of the individual patient. In general, the total daily dose range, is Kingdom Plantae, including terrestrial, aquatic or other from about 0.01 mg to about 2,000 mg of the plant extract(s) plants that can be subjected to the methodology described and/or active ingredient administered in about one to ten herein in order to generate an extract that can be tested for doses or applications. its ability to inhibited at least one of the above-listed skin EPs. 0188 Commercial Processes For Preparing Plant Extracts of the Invention 0196. Examples of potential plants include, but are not limited to, those belonging to the following classifications: 0189 The present invention contemplates the large-scale Superdivision Spermatophyta Seed plants; Division preparation of the plant extracts of the invention. The Coniferophyta Conifers; Class Pinopsida, Order Pinales: extracts can be prepared on a commercial scale using the Family Araucariaceae Araucaria family; Family Cephalo extraction process employed in the analytical scale prepa taxaceae Plum Yew family; Family Cupressaceae Cy ration the extract of interest. One embodiment of this aspect press family; Family Pinaceae Pine family; Family of the invention is presented in FIG. 3. In this embodiment, Podocarpaceae-Podocarpus family. Family Taxodi the Small-scale extraction procedure is simply scaled-up and aceae Redwood family; Order Taxales, Family Tax additional steps of quality control are included to ensure aceae Yew family: Division Cycadophyta Cycads, Class reproducible results. Similarly the process outlined in FIG. Cycadopsida, Order Cycadales, Family Cycadaceae— 5 can be adapted for scale-up for commercial purposes. Cycad family; Family Zamiaceae—Sago-palm family; Divi 0190. Also contemplated by the present invention are sion Ginkgophyta-Ginkgo, Class Ginkgoopsida, Order modifications to the Small-scale procedure that may be Ginkgoales, Family Ginkgoaceae-Ginkgo family: Division required during scale-up for industrial level production of Gnetophyta Mormon tea and other gnetophytes, Class the extract. Such modifications include, for example, alter Gnetopsida, Order Ephedrales, Family Ephedraceae Mor ations to the solvent being used or to the extraction proce mon-tea family; Order Gnetales, Family Gnetaceae—Gne dure employed in order to compensate for variations that tum family: Division Magnoliophyta-Flowering plants, occur during scale-up and render the overall procedure more Class Liliopsida Monocotyledons, Subclass Alismatidae, Order Alismatales, Family Alismataceae Water-plantain amenable to industrial scale production, or more cost effec family, Family Butomaceae Flowering Rush family, Fam tive. Modifications of this type are standard in the industry ily Limnocharitaceae Water-poppy family; Order Hydro and would be readily apparent to those skilled in the art. charitales, Family Hydrocharitaceae Tape-grass family; 0191 Process for Identifying Additional Plant Extracts Order Najadales, Family Appnogetonaceae—Cape-pond weed family, Family Cymodoceaceae—Manatee-grass fam 0192 The present invention further provides for a rapid ily, Family Juncaginaceae—Arrow-grass family, Family method for screening plant extracts to identify those capable Najadaceae Water-nymph family, Family Posidoni of inhibiting one or more of MMP-1, MMP-2, MMP-3, aceae—Posidonia family, Family Potamogetonaceae— MMP-9 and HLE, which are suitable for incorporation into Pondweed family, Family Ruppiaceae-Ditch-grass family, the dermatological formulations of the invention. Family Scheuchzeriaceae Scheuchzeria family, Family 0193 The process comprises the following general steps: Zannichelliaceae Homed pondweed family, Family (a) generating a plurality of extracts from plant material by Zosteraceae—Eel-grass family; Subclass Arecidae, Order Solvent extraction; (d) analysing the ability of each plant Arales, Family Acoraceae-Calamus family, Family extract to inhibit one or more of MMP-1, MMP-2, MMP-3, Araceae Arun family, Family Lemnaceae-Duckweed MMP-9 and HLE; and selecting those extracts that are family; Order Arecales, Family Arecaceae Palm family; capable of inhibiting the activity of at least one of the listed Order Cyclanthales, Family Cyclanthaceae Panama Hat EPs. The extracts exhibiting inhibitory activity can then be family; Order Pandanales, Family Pandanaceae Screw screened for their ability to affect one or more cellular pine family; Subclass Commelinidae, Order Commelinales, activities in skin cells, such as attenuating the breakdown of Family Commelinaceae Spiderwort family, Family Maya a structural component of the ECM (i.e. collagen, fibronec caceae—Mayaca family, Family Xyridaceae Yellow-eyed tin, fibrillin and/or elastin); attenuating endothelial cell Grass family; Order Cyperales, Family Cyperaceae Sedge migration; increasing collagen production; attenuating UV family, Family Poaceae Grass family; Order Eriocaulales, induced extracellular protease activity and attenuating trac Family Eriocaulaceae Pipewort family; Order Juncales, tional forces generated by fibroblasts. Those extracts that are Family Juncaceae Rush family; Order Restionales, Family effective in the cellular screen are considered to be suitable Joinvilleaceae Joinvillea family; Order Typhales, Family candidates for inclusion in the dermatological formulations Sparganiaceae—Bur-reed family, Family Typhaceae—Cat provided that they exhibit suitable stability and toxicity tail family: Subclass Liliidae, Order Liliales, Family Aga profiles. vaceae Century-plant family, Family Aloeaceae Aloe family, Family Dioscoreaceae Yam family, Family Hae 0194 The plurality of extracts in step-(a) above can be modoraceae—Bloodwort family, Family Hanguanaceae— generated from plant-material from a single plant Source Hanguana family, Family Iridaceae—Iris family, Family using different solvents or the plurality of extracts can be Liliaceae Lily family, Family Philydraceae Philydraceae generated by first selecting a group of plants of interest, family, Family Pontederiaceae Water-Hyacinth family, harvesting plant material from each plant in the group, then Family Smilacaceae Catbrier family, Family Stemo extracting the plant material with a solvent or solvents to naceae Stemona family, Family Taccaceae-Tacca family; generate a plurality of extracts. Order Orchidales, Family Burmanniaceae Burmannia US 2007/01 22492 A1 May 31, 2007 family, Family Orchidaceae Orchid family; Subclass Zin Family Monotropaceae Indian Pipe family, Family Pyro giberidae, Order Bromeliales, Family Bromeliaceae Bro laceae Shinleaf family; Order Lecythidales, Family meliad family; Order Zingiberales, Family Cannaceae— Lecythidaceae—Brazil-nut family; Order Malvales, Family Canna family, Family Costaceae—Costus family, Family Bombacaceae—Kapok-tree family, Family Elaeocar Heliconiaceae—Heliconia family, Family Marantaceae— paceae—Elaeocarpus family, Family Malvaceae—Mallow Prayer-Plant family, Family Musaceae Banana family, family, Family Sterculiaceae Cacao family, Family Tili Family Zingiberaceae-Ginger family: Class Magnoliop aceae—Linden family; Order Nepenthales, Family Droser sida Dicotyledons, Subclass Asteridae, Order Asterales, aceae—Sundew family, Family Nepenthaceae—East Indian Family Asteraceae Aster family; Order Callitrichales, Pitcher-plant family, Family Sarraceniaceae Pitcher-plant Family Callitrichaceae Water-starwort family, Family family; Order Primulales, Family Myrsinaceae Myrsine Hippuridaceae Mare's-tail family; Order Calycerales, family, Family Primnulaceae Primrose family, Family Family Calyceraceae—Calycera family; Order Campanula Theophrastaceae. Theophrasta family; Order Salicales, les, Family Camnpanulaceae F Bellflower family, Family Family Salicaceae Willow family; Order Theales, Family Goodeniaceae—Goodenia family, Family Spheno Actinidiaceae Chinese Gooseberry family, Family Caryo cleaceae—Spenoclea family; Order Dipsacales, Family caraceae-Souari family, Family Clusiaceae—Mangosteen Adoxaceae—Moschatel family, Family Caprifoliaceae— family, Family Dipterocarpaceae Meranti family, Family Honeysuckle family, Family Dipsacaceae Teasel family, Elatinaceae Waterwort family, Family Marcgraviaceae— Family Valerianaceae—Valerian family; Order Gentianales, Shingle Plant family, Family Ochnaceae Ochna family, Family Apocynaceae—Dogbane family, Family Asclepia Family Theaceae Tea family; Order Violales, Family daceae-Milkweed family, Family Gentianaceae—Gentian Begoniaceae—Begonia family, Family Bixaceae—Lipstick family, Family Loganiaceae—Logania family; Order Lami tree family, Family Caricaceae-Papaya family, Famnily ales, Family Boraginaceae-Borage family, Family Lami Cistaceae—Rock-rose family, Family Cucurbitaceae-Cu aceae—Mint family, Family Lennoaceae—Lennoa family, cumber family, Family Datiscaceae—Datisca family, Fam Family Verbenaceae Verbena family, Order Plantaginales, ily Flacourtiaceae Flacourtia family, Family Frankeni Famnily Plantaginaceae Plantain family; Order Rubiales, aceae—Frankenia family, Family Loasaceae—Loasa Family Rubiaceae Madder family; Order Scrophulariales, family, Family Passifloraceae Passion-flower family, Fam Family Acanthaceae Acanthus family, Family Bignoni ily Tamaricaceae—Tamarix family, Family Turneraceae— aceae Trumpet-creeper family, Family Buddleiaceae— Turnera family, Family Violaceae Violet family; Subclass Butterfly-bush family, Family Gesneriaceae Gesneriad Hamamelidae, Order Casuarinales, Family Casuarinaceae— family, Family Lentibulariaceae Bladderwort family, Fam She-oak family; Order Fagales, Family Betulaceae—Birch ily Myoporaceae—Myoporum family, Family Oleaceae— family, Family Fagaceae Beech family; Order Hamamel Olive family, Family Orobanchaceae Broom-rape family, idales, Family Cercidiphyllaceae-Katsura-tree family, Family Pedaliaceae Sesame family, Family Scrophulari Family Hamamelidaceae Witch-hazel family, Family Pla aceae Figwort family; Order Solanales, Family Convolvu tanaceae-Plane-tree family; Order Juglandales, Family. laceae—Morning-glory family, Family Cuscutaceae—Dod Juglandaceae Walnut family; Order Leitneriales, Family der family, Family Fouquieriaceae Ocohillo family, Leitneriaceae Corkwood family; Order Myricales, Family Family Hydrophyllaceae Waterleaf family, Family Men Myricaceae Bayberry family; Order Urticales, Family yanthaceae—Buckbean family, Family Polemoniaceae— Cannabaceae-Hemp family, Family Cecropiaceae—Ce Phlox family, Family Solanaceae Potato family: Subclass cropia family, Family Moraceae Mulberry family, Family Caryophyllidae, Order Caryophyllales, Family Achatocar Uhlaceae Elm family, Family Urticaceae Nettle family; paceae Achatocarpus family, Family Aizoaceae—Fig Subclass Magnoliidae, Order Aristolochiales, Family Aris marigold family, Family Amaranthaceae—Amaranth family, tolochiaceae Birthwort family; Order Illiciales, Family Family Basellaceae—Basella family, Family Cactaceae— Illiciaceae—Star-anise family, Family Schisandraceae— Cactus family, Family Caryophyllaceae Pink family, Fam Schisandra family; Order Laurales, Family Calycan ily Chenopodiaceae Goosefoot family, Family Mollugi thaceae Strawberry-shrub family, Family Hemandi naceae—Carpet-weed family, Family Nyctaginaceae—Four aceae—Hemandia family, Family Lauraceae—Laurel o'clock family, Family Phytolaccaceac—Pokeweed family, famnily, Family Monimiaceae Monimia family; Order Family Portulacaceae Purslane family; Order Plumbagi Magnoliales, Family Annonaceae-Custard-apple family, males, Family Plumbaginaceae Leadwort family; Order Family Canellaceae Canella family, Family Magnoli Polygonales, Family Polygonaceae—Buckwheat family; aceae—Magnolia family, Family Myristicaceae Nutmeg Subclass Dilleniidae, Order Batales, Family Bataceae— family, Family Sonneratiaceae Sonneratia family, Family Saltwort family; Order Capparales, Family Brassicaceae— Winteraceae Wintera family; Order Nymphaeales, Family Mustard family, Family Capparaceae—Caper family, Fam Cabombaceae Water-shield family, Family Ceratophyl ily Moringaceae Horse-radish tree family, Family laceae Hornwort family, Family Nelumbonaceae Lotus Resedaceae Mignonette family; Order Diapensiales, Fam lily family, Family Nymphaeaceae Water-lily family; ily Diapensiaceae Diapensia family; Order Dilleniales, Order Papaverales, Family Fumariaceae Fumitory family, Family Dilleniaceae Dillenia family, Family Paeoni Family Papaveraceae Poppy family; Order Piperales, aceae Peony family; Order Ebenales, Family Ebenaceae— Family Chloranthaceae Chloranthus family, Family Pip Ebony family, F Family Sapotaceae Sapodilla family, eraceae—Pepper family, Family Saururaceae—Lizard's-tail Family Styracaceae Storax family, Family Symplo family; Order Ranunculales, Family Berberidaceae Bar caceae Sweetleaf family; Order Ericales, Family Clethra berry family, Family Lardizabalaceae—Lardizabala family, ceae—Clethra family, Family Cyrillaceae—Cyrilla family, Family Menispermaceae Moonseed family, Family Family Empetraceae Crowberry family, Family Epacri Ranunculaceae—Buttercup family, Family Sabiaceae Sa daceae—Epacris family, Family Ericaceae—Heath family, bia family: Subclass Rosidae, Order Apiales, Family Api US 2007/01 22492 A1 May 31, 2007

aceae—Carrot family, Family Araliaceae-Ginseng family; Aconitum spp., Acorus calamus, Actaea racemosa, Actinidi Order Celastrales, Family Aquifoliaceae Holly family, colonicta, Actinidia arguta, Actinidia chinensis, Actinidia Family Celastraceae Bittersweet family, Family Coryno colonicta, Adansonia digitata, Adianthum radiatum, Adian carpaceae-Karaka family, Family Hippocrateaceae— thum trapezieformis, Adiantum pedatum, Adiantum tenerum, Hippocratea family, Family Icacinaceae—Icacina family, Aechmea luddemoniana, Aesculus hypocastanum, Aesculus Family Stackhousiaceae Stackhousia family; Order waertilensis, Aesculus woerlitzenis, Aessopteria crasifolia, Comales, Family Comaceae—Dogwood family, Family Aframomum melegueta, Agaricus bisporus, Agastache foe Garryaceae Silk Tassel family, Family Nyssaceae Sour niculum, Agastache mexuicana, Agatis robusta, Ageratum Gum family; Order Euphorbiales, Family Buxaceae Box conizoides, Aglaonema commutatus, Agrimonia eupatora, wood family, Family Euphorbiaceae—Spurge family, Fam Agropyron Cristatum, Agropyron repens, Agrostis alba, ily Simnmondsiaceae Jojoba family; Order Fabales, Fam ily Fabaceae Pea family; Order Geraniales, Family Agrostis stolonifera, Ailantus altissima, Ajuga reptans, Balsaminaceae Touch-me-not family, Family Gerani Alcea rosea, Alchemilla mollis, Alchemilla sp., Alium cer aceae—Geranium family, Family Limnanthaceae— mum, Alkanna tinctoria, Allium ampeloprasum, Allium Meadow-Foam family, Family Oxalidaceae Wood-Sorrel cepa, Allium fistulosum, Allium grande, Allium nutans, family, Family Tropaeolaceae—Nasturtium family; Order Allium porrum, Alium sativum, Allum Schoenoprasum, Haloragales, Family Gunneraceae-Gunnera family, Fam Albium sp., Allum tuberosum, Allium victorialis, Aloe vera, ily Haloragaceae Water Milfoil family; Order Linales Alpinia officinarum, Althaea officinalis, Alum japonica, Family Erythroxylaceae-Coca family, Family Linaceae— Amaranthus caudatus, Amaranthus retroflexus, Amaranthus Flax family; Order Myrtales, Family Combretaceae Indian tricolor; Ambrosia artemisiifolia, Amelanchier alnifolia, Almond family, Family Lythraceae—Loosestrife family, Amelanchier Canadensis, Amelanchier sanguinea, Amelan Family Melastomataceae Melastome family, Family chier sanguineaxA. laevis, Amelanchier spicata, Amigdalus Myrtaceae-Myrtle family, Family Onagraceae—Evening nana, Amsonia tabenaemontana, Ananas comosus, Anapha Primrose family, Family Punicaceae Pomegranate family, lis margaritacea, Anemona japonica, Anethum graveolens, Family Thymelaeaceae Mezereum family, Family Tra Angelica archangelica, Angelica dahurica, Angelica sinen paceae Water Chestnut family; Order Podostemales, Fam sis, Antericum ramosum, Anthemis tinctoria, Anthoxanthum ily Podosteimaceae River-weed family; Order Polygala Odoratum, Anthriscus cerefolium, Anthurium altersianum, les, Family Krameriaceae—Krameria family, Family Anthurium andreanum, Anthurium elegans, Anthurium Malpighiaceae Barbados Chemy family, Family Polygal guildingii, Anthurium hookeri, Anthurium magnificum, aceae Milkwort family; Order Proteales, Family Pro Anthyrium filis-femina, Anthyrium nopponicum, Apium gra teaceae Protea family; Order Rafflesiales, Family Raffle veolens, Apocynum cannabinum, Arachis hypogaea, Aralia siaceae Rafflesia family; Order Rhamnales, Family cordata, Aralia nudicaulis, Aralis mandshurica, Archirantus Elaeagnaceae—Oleaster family, Family Rhamnaceae— bidentata, Arctium lappa, Arctium minus, Arctostaphylos Buckthorn family, Family Vitaceae Grape family; Order uva-ursi, Armoracea rusticana, Armoraica ristica, Aronia Rhizophorales, Family Rhizophoraceae Red Mangrove melanocarpa, Aroniaxprunifolia, Arrhenatherum elatius, family; Order Rosales, Family Brunelliaceae—Brunellia Artemisia abrotanum, Artemisia absinthium, Artemisia dra family, Family Chrysobalanaceae-Cocoa-plum family, cunculus, Artemisia ludoviciana, Artemisia vulgaris, Family Connaraceae—Cannarus family, Family Crassu Asarum europaeum, Asclepias incamata, Asclepias tube laceae Stonecrop family, Family Crossosomataceae— rosa, Asimina triloba, Asorum Canadensis, Asparagus offi Crossosoma family, Family Cunoniaceae—Cunonia family, cinalis, Asplenium australasicum, Aster spp., Aster-Nova Family Grossulariaceae Currant family, Family anglicae, Astilbe chinensis, Astilbexarendsii, Astillboides Hydrangeaceae—Hydrangea family, Family Pittospora tabularis, Astragulus sinicus, Athyrium asperum, Atriplex ceae—Pittosporum family Family Rosaceae—Rose family, hortensis, Atropa belladonna, Austolachia australis, Avena Family Saxifragaceae-Saxifrage family, Family Surian sativa, Averrhoa carambola, Bactisia australis, Baptisia aceae Suriana family; Order Santalales, Family Balano tinctoria, Barbaric sp., Beckmannia eruciformis, Begonia phoraceae—Balanophora family, Family Eremolepi convolvulacea, Begonia eminii, Begonia glabra, Begonia daceae—Catkin-mistletoe family, Family Loranthaceae— mannii, Begonia polygonoides, Bellis perennis, Berberis Showy Mistletoe family, Family Olacaceae Olax family, thungergi, Berberis vulgaris, Bergenia crassifolia, Berge Family Santalaceae Sandalwood family, Family Vis niaxschmidtii, Beta vulgaris, Betula alba, Betula caceae Christmas Mistletoe family; Order Sapindales, alleghaniensis, Betula daurica, Betula glandulosa, Betula Family Aceraceae—Maple family, Family Anacardiaceae— nigra, Betula pendula, Bocconia Cordata, Boechimeria Sumac family, Family Burseraceae—Frankincense family, boloba, Boesenbergia rotunda, Boletus edulis, Borago offi Family Hippocastanaceae-Horse-chestnut family, Family cinalis, Boxus sempervirens, Brassica cepticepa, Brassica Meliaceae—Mahogany family, Family Rutaceae—Rue fam chinensis, Brassica juncea, Brassica napa, Brassica napus, ily, Family Sapirdaceae Soapberry family, Family Sima Brassica nigra, Brassica oleracea, Brassica rapa, Bromelia roubaceae—Ouassia family, Family Staphyleaceae—Blad balansae, Bromus inermis, Brugmansi graveolens (ralf), derniut family, Family Zygophyllaceae—Creosote-bush Brugmansia suaveolens, Bruginansia suaveolens, Buddleja family. davidii, Bupleurum falcatum, Butomus umbellatus, Buxus microphilla japonica'. Buxus microphylla, Cachris alpina, 0197). In one embodiment, potential plants comprise: Cactus officinalis, Caladium spp., Calamagrostis arundi Abelmoschus esculentus, Abies balsamea, Abies cepha flora, Calamintha nepeta, Calathea zebrina, Calendula offi lonica, Abies firma, Abies lasiocarpa, Acer campestre, Acer cinalis, Calicatus floridus, Camelia sinensis, Campanula mandshurica, Acer palmaturn "burgundy.' Acer tataricum, carpatica, Campanula rapunculus, Canna indica, Acer truncatum, Achillea millefolium, Achillea ptarmica, Cantharellus cibarius, Capparis spinosa inemis, Capsella Achillea tomentosa, Acolypha hispida, Aconitum napellus, bursa-pastoris, Capsicum annuum, Capsicum frutescens, US 2007/01 22492 A1 May 31, 2007 20

Carex morrowii, Carica papaya, Carlina acaulis, Carpinus Fuchsia magellanica, Fuchsia spp., Fucus vesiculosus, caroliniana, Carthamus tinctorius, Carum capsicum, Carum Fumaria officinalis, Galinsoga quadriradiata, Galium carvi, Carya cordiformis, Caryota ureus, Casia hebecarpa, aparine, Galium odoratum, Gallium sporium, Gardenia Castanea sativa, Castanea spp., Celosia cristata, Celtis jasminoides, Gaultheria hispidula, Gaultheria procumbens, occidentalis, Centaurea dealbata, Centaurea soilstitialis, Genista multibracteata, Gentiana cruciata, Gentiana lit Centauria maculata, Cerastium tomentosum, Cerasus torala, Gentiana lutea, Gentiana macrophylla, Gentiana japonica, Cerasus maghabab, Ceratoramia mexicana, tibetica, Geranium maculata, Geranium phaeum, Geranium Chaenomelesxsuperba, Chaernomelis superba, Chaero pratense, Geranium sanguineum, Geraniumxcant phyllum bulbosum, Chamaemelum nobile, Charnaechrista abrigiense, Geum fanieri, Geum macrophyllum, Geum fasciculata, Charnaeciparis pisifera, Chelidonium majus, rivale, Gingko biloba, Glaux maritima, Glechoma hedera Chenopodium album, Chenopodium bonus-henricus, Che cea, Glyceria maxima, Glycine max, Glycyrrhiza glabra, nopodium quinoa, Chrysanthemum coronarium, Cicer ari Gnetum guemon, Gossypium herbaceum, Gratiola officina etinum, Cichorium endivia Subsp. endivia, Cichorium inty lis, Gravilea robusta, Guizotia abyssinica, Haemanthus bus, Cinnamomum verum, Cirsium arvense, Cissus discolor, katharina, Hamamelis mollis, Hamamelis virginiana, Haser Cistus incanus, Citinis coggriaria, Citrullus colocynthis, trilobum, Hedeoma pullegioides, Hedychium coronarium, Citrullus lanatus, Citrus limettoides, Citrus limon, Citrus Hedychium spp., Helenium spp., Helianthus annus, Helian reticulata, Citrus sinensis, Citrusxparadisi, Clematis thus stumosus, Helianthus tuberosus, Helichrysum angusti alpina, Clematis armandii, Clematis chiisanensis, Clematis folium, Helichrysum thians chanicum, Heliotropium arbore rectae, Clerodendrurn speciossicum, Cobiaeum varillartum, scens, Helleborus niger; Heraclelum pubescens, Herba Coccoloba Caracasana, Cocculus laurifolius, Cocos schizonepetae, Hemerocalis spp., Hibiscus cannabinus, nucifera, Coix lacryma jobi, Colocasia spp., Comus mass, Hissopus zeraucharicus, Hiuga reptans, Hordeum hexasti Convalaria majalis, Conyza-Canadensis, Corchorus Olito chon, Hordeum vulgare, Hordeum vulgare subsp. vulgare, rius, Coreopsis verticillata, Coriandruim sativum, Cornus Hosta fortuna, Hosta fortunaea, Hosta lancefolia, Hosta alba, Cornus Canadensis, Cornus mas, Cornus sericea, Sieboldiana, Hosta zibalda, Houttuynia cordata, Humulus Coronola varia, Corylus avelana, Corylus maxima, Cos lupulus, Hydrangea quercifolia, Hydrastis Canadensis, mos Sulphureus, Cotinus Coggy'gria, Cotoneaster fangianus, Hydrocotile asiatica, Hylotelephium spp., Hymenoxys Cotoneaster horisontalis, Cotynus Cog'gria, Crambe cordi hoopesi, Hyoscyamus niger, Hypericum henryi, Hypericum folia, Cramble cardifolia, Crataegus praegophyrum, Cra perforatum, Hypericum spp., Hypomyces lactifluorum, Hip taegus sanguinea, Crataegus spp., Crataegus submollis, poach rhamnoides, Hyssopus officinalis, Iberis amara, Crategus macrophyllum, Crithmum maritimum, Cryptotae Iberis sempervirens, Ilex agnifolium, Ilex comuta, Inula nia Canadensis, Crytomium fortunei, Cucumis anguria, helenium, Ipomea tricolor, Ipomoea aquatica, Ipomoea Cucumis melo, Cucumis metuliferus, Cucumis sativus, batatas, Iris alida, Iris pseudocarpus, Iris versicolor; Isatis Cucurbita maxima, Cucurbita moschata, Cucurbita pepo, tinctoria, Jacobinia sp., Jasminum frutocarus, Jeffersonia Cullen corylifolium, Cuminum cyminum, Cupress lusitanica, diphylla, Juca sp., Juglands regia, Juglans migra, Juniperus Cupressus sempervirens, Curcuna longa, Curcuma “blue pacific'. Juniperus communis, Kevleiteria paniculata, Zedoaria, Cycas cirinalis, Cyclonia Oblonga, Cymbopogon Kochia scoparia, Koeleria glauca, Kolkwitzia amabilis, citratus, Cymbopogon martinii, Cynara cardunculus Subsp. Korria japonica, Krameria lappacea, Lactuca sativa, Lac cardunculus, Cynnamomum zeylonicum, Cyperus alternifo tuca serriola, Lal lab purpurea, Lamiastrum galeobdolon, lius, Cyperus esculentus, Dactylis glomerata, Dahlia spp., Lapia dulcis, Laportea Canadensis, Larix dedidua, Laserpi Darura stramonium, Datisca cannabina, Datura metel, tium latifolium, Lathyrus sativus, Lathyrus Sylvestris, Lau Datura stramonium, Daucus carota, Deutria scabra, Dief rus nobilis, Lavandula angustifolia, Lavandula latifolia, fenbachia leopoldii, Diefenbachia segiunae, Digitalis lutea, Lavandula officinalis, Ledum groenlandicum, Lens culinaris Digitalis purpurea, Dimocarpus longan, Diopiros kaka, Subsp. culinaris, Lentinus edodes, Leontopodium alpinum, Dioscorea batatas, Diospyros kaki, Dipsacus sativus, Dirca Leonurus Cardiaca, Lepidium sativum, Leucanthemum vul palustris, Dolichos lablab, Dracaena fragrams, Dracaena gare, Levisticurn officinale, Liatris spinata, Liclun barba sp., Dryopteris filis-max, Dryopteris filix-mas, Echinacea tum, Ligularia dentata, Ligustrum vulgare, Linaria vulgaris, purpurea, Echinochloa frumentacea, Echinops sphae, Elea Lindera benzoin, Linium hirsutum, Linum usitatissimum, gnus angustifolia, Eleagnus cemutata, Eleusine Coracana, Lippa dulcis, Litchi chinensis, Livistona fragrams, Lobelia Encephalaris horridum, Epilobium augustifolium, Equise siphitica, Lolium multiflorum, Lolium perenne, Lonicera tum hyemale, Equisetum variegatum, Erigeron speciosus, ramosissima, Lonicera syringantha, Lotus cornicuiatus, Eriobotria japonica, Eriobotrya japonica, Eruca vesicaria, Lotus tetragonolobus, Luglands nigra, Lunaria annua, Erungiurm campestre, Erysimumw perofskianum, Eryth Lupinus luteaus, Lupinus polyphyllus, Luzula Sylvatica, rinia caffra, Erythrinia Crista, Erythrinia glabeliferus, Lychnis chalcedonica, Lycodium japonicum, Lycopersicon Eschscholzia Californica, Eucaliptus rudis, Eucomia uluri esculentum, Lycopersicon pimpinellifolium, Lysimachia folia, EuOnimus elata, Euonomus europea, Euonomus ver clethroides, Lythrum salicaria, Madia sativa, Magnolia rucosa, Euphorbia amygdaloides, Fagopyrum esculentum, agrifolia, Magnolia cobus, Magnolia loebheril, Magnolia Fagopyrum suffruticosum, Fagopyrum tataricum, Fagus sil Stellata, Magnoliaxloebneri, Malus hupehensis, Malus Vatica, Fautenousus qualiqualia, Festuca rubra, Feucrium prunifolia, Malus spp., Malva moschata, Malva Sylvestris, hamedris, Ficus benjamini, Ficus elastica, Ficus purnila, Malva verticillata, Mangifera indica, Manihot esculenta, Ficus religiosa, Ficus sp., Ficus triangularis, Filipendula Marrubium vulgare, Matricaria recutita, Matricaria spp., rubra, Filipendula ulmaria, Filipendula vulgaris, Foenicu Matteuccia pensylvanica, Matteucia strutioptoris, Medi lum vulgare, Foenix Zeulonica, Forsithsia suspensa, Forsit cago sativa, Melaleuca altenifolia, Melilotus albus, Melilo sia europea, Forsythiaxintermedia, Fortunella spp., tus officinalis, Melissa officinalis, Mentha arvensis, Mentha Fragariaxananassa, Frangula alnus, Fraxinus excelsior, pullegium, Mentha spicata, Mentha suaveolens, Menthax US 2007/01 22492 A1 May 31, 2007 piperita, Menyanthes trifoliata, Mespilus germanica, imbricaria, Ouercus migra, Quercus robur “fastigiata,'Oue Metasequoia glyptotrobioldes, Metrosideros excelsa, Micro rcus rubra, Quercus trojana, Raphanus raphanistrum, biata decussata, Microlepia platphylla, Microlepia platy Raphanus sativus, Ratibiunda columnus-Fera, Rauwolfia phylla, Microsorium punctatum, Minispermum dauricum, tetraphylla, Rehmannia glutinosa, Reseda luteola, Reseda Mirica certifera, Miscanthus sacchariflorus, Miscanthus Odorata, Rheum officinale, Rheum palmatum, Rheumxhybri Sinensis, Momordica charantia, Monarda didyma, Monarda dum, Rhododendron spp., Rhus aromatica, Rhus toxico fistulosa, Monarda spp., Monstera deliciosa, Monstera per denta, Rhus trilobata, Ribes americanum, Ribes grossularia, tusa, Montia perfoliata, Morms alba, Murraya exotica, Ribes nigrum, Ribes Sylvestre, Ribes uva-crispa, Ribes x Musa textilis, Musaxparadisiaca, Myrica pensylvanica, nidigrolaria, Ricinus communis, Rimula japonica, Rodger Myrthus communis, Nasturtium officinale, Nepeta cataria, sia podophylla, Rodgersia spp., Rosa cocanica, Rosa mul Nicodernia diversifolia, Nicotiana rustica, Nicbtiana tiflora, Rosa rugosa, Rosmriarinus officinalis, Rubus tabacum, Nigella sativa, Ocimirnum Basilicum, Ocirnum allegheniensis, Rubus arcticus, Rubus Canadensis, Rubus tenuiflorum, Oenothera biennis, Oenothera fruticosa subsp idaeus, Rubus occidentalis, Rubus phoenicolasius, Rubus fruticosa, Olea europaea, Olea Olcaster, Onobrychis vicii pubescens, Rubus thibetanus, Rudbeckia maxima, Rumex folia, Onoclea sensibilis, Ophiopogon japonicus, Opuntia acetosa, Rumex acetosella, Rumex crispus, Rumex patientia, spp., Oreopanax capitata, Origanum majorana, Origanum Rumex scutatus, Ruschia indurata, Ruta graveolens, Sac vulgare, Oryza sativa, Osmanthus spp., Osmunda regalis, charum officinarum, Salis babilonics, Salix purpurea, Salix Osmundastrum claytonionum, Ostrea carpinifolia, Ostrea tamarisifolia, Salvia elegans, Salvia officinalis, Salvia connote, Oxalis deppei, Oxobachus nictogenea, Oxyria Sclarea, Salvia Sylvestris, Sambucus Canadensis, Sambucus digyna, Pachyra affinis, Paeonia daurica, Paeonia lacti ebulus, Sambucus migra, Sanchezia nobilis, Sanguisorba flora, Paeonia rubra, Paeonia spp., Paeonua suffructicisa, minor, Sanguisorba officinalis, Santolina chamnaecyparis Panax quinquefolius, Panicum miliaceum, Parrotia persica, sus, Saponaria officinalis, Satureja hortensis, Satureja mon Parthenosicus tricuspidata, Passiflora caerulea, Passiflora tana, Satureja repandra, Schisandra chinensis, Scolymus spp., Pastinaca sativa, Pegamun hamalis, Pelargonium Zon hispanicus, Scorzonera hispanica, Scotch pine, Scrophu ale, Pennisetum alopecuroides, Penstemon digitalis, Penta laria nodosa, Scutellaria certicola, Scutellaria lateriflora, phylloides fruticosa, Perilla frutescens, Persea americana, Scutellarian altissirna, Secale cereale, Sechium edule, Petasites japonicus, Petroselinum crispum, Peucedanum Sedum alburn, Sedum telchium, Sempervivum tectorum, cervaria, Peucedanum Oreaselinum, Pfaffia paniculata, Senecio platifila, Senecio vuigaris, Senseviera sp., Serenoa Phacelia tanacetifolia, Phalaris arundinacea, Phalaris repens, Seringa josiceae, Serratula tinctoria, Seruginea canariensis, Phaseolus acutifolius, Phaseolus coccineus, uffruticisa, Sesamum indicum, Sesbania exaltata, Sesbania Phaseolus vulgaris, Phebodium aureum, Philadelphus coro speciosa, Setaria italica, ibirea altaiensis, Sidalcea spp., narius, Philodendron amurense, Phleum pratense, Phlox Silene vulgaris, Silybum marianum, Sinapis alba subsp. paniculata, Phoenix dactylifera, Phylidendron speciosus, alba, Siringa vulgaris, Sium sisarum, Sluffera sp., Solanum Phyllanthus grandifolium, Phyllitis scolopendrium, Phyrna duicamara, Solanum melongena, Solanum scabrum, tosorus scolopendria, Physalis alkekengi, Physalis Creti Solanum tuberosum, Soleirolia Soleirolii, Solidago caesia, cola, Physalis grisea, Physalis philadelphica, Physalis spp., Solidago Canadensis, Solidago spp., Solidago virgaurea, Physostegia virginiana, Phytolacca americana, Picea Solidagoxhybrida, Sonchus oleraceus, Sorbocotoneaster Schrenkiana, Pieras japonica, Pigelia pennata, Pimpinella sp., Sorbus aucuparia, Sorbus cominicta, Sorghum bicolor, anisum, Pinus bungiana, Pinus Cembra, Pinus mugo, Pinus Sorghumx.drummondii, Spartina potentiflora, Spathiphyllum pinea, Pinus punila, Pinus Salinifolia, Pinus silvestris, cochlearispaturn, Spathiphyllum grandiflorum, Spinacia Pinus sirtrobus, Pinus strobus, Piper chaba, Piper nigrum, oleracea, Stachis lanata, Stachys affinis, Stachys byzantina, Pisum sativum, Pithecelobium unguis, Pittisporum tibica, Stachys macrantha, Staphylea trifolia, Stellaria graminea, Plantago coronopus, Plantago major; Plantago minor, Pla Stellaria media, Stephanandra incisa, Stepochlaena tenui tanus acidentalis, Platicada grandiflora, Plectranthus fru folia, Sterulia elata, Stevartia coreana, Stewartia pseudoca ticosus, Plectranthus spp., Pleurotus spp., Plumbago zey melia, Stipa capillata, Strelitzia reginae, Sulda sanganea, lanica, Poa compressa, Poa pratensis, Podocarpus Sundapsis spp., Symphitium officinalis, Symphoricarpbs spinulosus, Podophyllum anodii, Podophyllum peltatum, albus, Symphoricarpos Orbiculatus, Symphytum officinale, Poligonum aviculare, Poligomun latifolia, Polygonium Odo Syngonium aurutum, Syngonium podophyllum, Taccus ratum, Polygonum aviculare, Polygonum chinense, bacata, Tagetes minuta, Talictrum minus, Talictrum sp., Polygonum cuspidatum, Polygonum pensylvanicum, Tamarindus india, Tamarindus indica, Tanacetum bal Polygonum persicaria, Polymonium ceruleum, Polyschium sanita, Tanacetum balsamita Subsp. balsamita, Tanacetum braunii, Pongamia pinnata, Pontederia Cordata, Populus cinerariifolium, Tanacetum parthenium, Tanacetum vulgare, incrassata, Populus tremula, Populusxpetrowskyana, Tapeinochilos spectabilis, Taraxacum officinale, Taraxacum Portulaca oleacea, Potentilla alba, Potentilla anserina, officinalis, Taxodium dixticum, Taxus cuspidata, Taxus hik Potentilla fruticosa, Poterium sangiusorba, Primula veris, sii, Taxus media, TaxusXmedia, Tetraclinis articulata hinen Princepia sp., Prunella vulgaris, Prunus armeniaca, Prunus sis, Tetradenia riparia, Teucrium chamaedrys, Thalictrum cerasifera, Prunus cerasus, Prunus persica, Prunus aquilegiifolium, Thalictumi flavum, Thlaspi arvense, Thuja serotica, Prunus spp., Prunus tomentosa, Prunus xocane, occidentalis, Thymus camosus, Thymus Cretaceus, Thymus Psathyrostachys juncea, Pseudotsuga menzisia, Psidium Cytridorus “aureus, Thymus fragantissimus, Thymus herba guajava, Psidium spp., Psychotria metbacteriodomasica, barona, Thymus lemabarona, Thymus portugalense, Thymus Psychotria nigropunctata, Pteridium aquilinum, Pterigota praecox, Thymus praecox. Subsp. arcticus, Thymus alata, Puansetia sp., Pulmonaria molissima, Pulmonaria pseudollamginosus, Thymus pseudolanuginosus, Thymus officinalis, Pulmonaria saccharata, Punica granatum, Pyrus pulleglodes "lemons'. Thymus pulliglodes, Thymus serphy communis, Pyrus pyrifolia, Quercus castanufolia, Quercus lum, Thymus speciosa, Thymus thrasicus, Thymus vulgaris, US 2007/01 22492 A1 May 31, 2007 22

Thymus vulgaris “argenteus.” Thymus vulgaris “oregano, Amsinckia intermedia; members of the Sunflower, Composi Thymus wooly, Thymusxcitriodorus, Tiarella cordifolia, tae, family, including Baccharis sarothroides, Monoptilon Tiarella spp., Tragopogon porrifolius, Tragopogon spp., belloides, Erieron divergens, Zinnia acerosa, Melampodium Trambe pontica, Trevesia Sungaica, Trichosanthes kirillowii, leucanthan, Chaenactis fremontii, Calycoseris Wrightii, Trifolium hybridum, Trifolium incaamatum, Trifolium pan Malacothrix Californica, Helianthus annus, H. niveus, Ger nomncum, Trifolium pratense, Trifolium repens, Trigonella aea canescens, Hymenothrix wislizenii, Encelia farinosa, foenum-graecum, Triticum aestivum, Triticum aestivum Psilostrophe cooperi, Baileya multiradiata, Bebbia juncea, Subsp. spelta, Triticum turgidium, TrolliusXcultorum, Tro Senecio douglasii, Trixis Californica, Machaeranthera teph paeolum majus, Tsuga Canadensis, Tsuga Canadensis rodes, Xvlorhiza tortifolia, Cirsinn neomexicanum, Anten “penola'. Tsuga diversifolia, Tsuga mertensiana, Tuja ori naria parviflora and Ch. douglasii; members of the caltrop, entalis “eligantissima, Tula Ocidentalis “columbia, Tulip Zygophyllaceae, family, including Larrea tridentata and tree, Tumera ulmifolia, Tissilago farfara, Tipha latifolia, Kallstroemia grandiflora; members of the mallow, Mal Ulmus americana, Ulmnus punila, Urtica dioica, Uschusa vaceae, family, including Hibiscus coulteri, H. denudatus sp., Uvywlaria perfoliata, Vaccinium angustifolium, Vac and Sphaeralcea ambigua; members of the phlox, Polemoni cinium corymbosum, Vaccinium macrocarpon, Valeriana aceae, family. Such as Luanthus aureus; members of the officinalis, Valerianella locusta, Veratrum nigrum, Veratnim unicorn plant, Martyniaceae, family, such as Proboscidiea viride, Verbascum thapsus, Verbena officinalis, Verium ole altheaefolia; members of the gourd, Cucurbitaceae, family, ander, Vemonia gigantea, Veronica austriaca ssp teucrium, Such as Cucurbita digitata; members of the lily, Lilaceae, Veronica beccabunga, Veronica officinalis, Viburnum opu family, including Calochoritus kennedy'i, DicheloStemma lus, Viburnum plicatum, Vicia faba, Vicia sativa, Vicia pulchellum, Allium macropetalum and Hesperocallis indu villosa, Vigna angularis, Vigna mungo, Vigna unguiculata, lata; members of the ocotillo, Fouquieriaceae, family, Vinca minor, Vincetocsicum officinalis, Vitis labrissa, Vitis including Fouquieria splendens; members of the figwort, spp., Weigela Coraeensis, Weigela hortensis, Withania son Scrophulariaceae, family, such as Castilleja sp., Penstemon nifera, XTriticosecale spp., Xanthium Sibiricum, Xanthium parryi and Orthocarpus purpurascens; members of the strumarium, Xanthosoma Sagittifolium, XeupressOcyparis acanthus, Acanthaceae, family, including Anisacanthus deylandii, Yucca elephantipes, Yucca filamentosa, Zea mays, thurberi, Justicia Californica and Ruellia nudiflora; mem Zelcova, Zingiber officinalis and Zingiber officinale. bers of the four o'clock, Nyctaginaceae, family. Such as Allionia incamata, Abronia villosa and Mirabilis multiflora; 0198 Groups of potential plants may also be selected members of the geranium, Geraniaceae, family, including based on their indigenous geographical regions. For Erodium cicutarium; members of the waterleaf, Hydrophyl example, one group of potential plants could comprise laceae, family, Such as Nama demissium, Phacelia bomby plants that are indigenous to arid regions, for example, those cina and Ph. distans; members of the bignonia, Bignoni located between 35° north latitude and 35° south latitude. In aceae, family, such as Chilopsis linearis; members of the accordance with another embodiment of the present inven Vervain, Verbenaceae, family, including Glandularia good tion, therefore, potential plants comprise: the agave, Aga dugii and Verbena neomexicana; members of the mint, vaceae, family including Such members as: Yucca elata, Y. Labiatae, family. Such as Hyptis emoryi and Salvia colum breviflora, Agave deserti, A. chrysantha, Dasylirion whee bariae; members of the broomrape, Orobanchaceae, family, leri; the buckwheat, Polygonaceae, family, such as Eri Such as Orobanche cooperi; members of the portulaca, ogonum fasciculatum; the crowfoot, Ranunculaceae, family, Portulaceae, family, such as Talinum auriantiacum; mem Such as Delphinium scaposum, Anemone tuberosa and D. bers of the carpet-weed, Aizoaceae, family. Such as Sesu parishii: the poppy, Papaveraceae, family, including Platys vium verrucosum: members of the flax, Linaceae, family, temon Californicus, Argemone pleiacantha, Corydalis aurea, Such as Linum lewisii; members of the potato, Solanaceae, Eschschoizia Californica and Ar. corymbosa; members of the family, including Nicotiana trigonophylla and Physalis mustard, Cruciferae, family, Such as Dithyrea Californica, Streptanthus carinatus and Lesquerella gordoni; members lobata; and members of the cochlospermum, Cochlosper of the legume, Leguminosae, family, Such as Acacia greggii, maceae, family, such as Amoreuxia palmatifida. Prosopis velutina, A. Constrica, Senna covesii, Cercidium 0199 If desired, the potential plant(s) can be subjected to floridum, C. microphyllum, Lotus huminstratus, Krameria a harvest stress treatment. A stress treatment comprises parvifolia, Parkinsonia aculeata, Calliendia eriophylla, contacting or treating the potential plant(s), or material from Lupinus arizonicus, Olyneya tesota, Astragalus lentigino the potential plant(s), with one or more stressor. The stressor sus, Psorothamunus spinosus and Lupinus sparsiflorus; can be a chemical compound or a physical treatment. members of the loasa family, Loasaceae, including Mentz Examples of suitable stressors are provided above. Various elia involucrata, M. pumila and Mohavea Confertifiora; combinations of stressors and treatment regimes can also be members of the cactus, Cactaceae, family, Such as Carn employed as would be apparent to one skilled in the art. egiea gigantia, Opuntia leptocaulis, Ferocactus wislizenii, 0200. The plant material may be used immediately after O. bigelovii, O. pheacantha, O. versicolor, O. filgida, harvest, or it can be stored for a period of time prior to Echinocereus engelmannii, Manmmillaria microcarpa, O. performing the extraction procedure(s). If desired, the plant basilaris, Stenocereins thurberi, O. violacea, M. tetrancis material can be treated prior to storage, for example, by tra, O. ramosissima, O. acanthocarpa, E. pectinatins and O. drying, freezing, lyophilising, or some combination thereof. arbuscula; members of the evening primrose, Onagraceae, Following treatment to prepare the plant material for stor family, Such as Oenothera deltoides, Camissonia clavifor age, the plant material may be stored for a period of time mis and Oe. primiveris; members of the milkweed, Ascl prior to preparation of the extract. The storage time may be epiadaceae, family, including Asclepias erosa, A. sublata of various duration, for example, the storage period may be and Sarcostemma cynanchoides; members of the borage, between a few days and a few years. In one embodiment of Boraginaceae, family, Such as Cryptantha augustifolia and the invention, the plant material is stored for a period of less US 2007/01 22492 A1 May 31, 2007

than one week. In another embodiment, the plant material is Surface area is presented to the solvent. For example, the stored for a period between one week to one month. In a plant material can be crushed or sliced mechanically, using further embodiment, the plant material is stored for a period a grinder or other device to fragment the plant parts into of between one month to six months. In other embodiments, Small pieces or particles, or the plant material can be frozen the plant material is stored for periods of between four liquid nitrogen and then crushed or fragmented into Smaller months to one year and for a period over one year in p1eces. duration. 0208. The solvent used for each extraction process can be 0201 The Extraction Process aqueous, alcoholic or organic, or a combination thereof. In 0202 Various extraction processes are known in the art one embodiment of the present invention, plant material is and can be employed in the process of the present invention extracted with an aqueous solvent. In another embodiment, (see, for example, International Patent Application WO an aqueous solvent comprising an aqueous buffer at pH 6-8 for a period of between 30 minutes to 8 hours at a tempera 02/06992). ture between about 4 to about 50° C. is used for the 0203. In one embodiment of the present invention the extraction. plant material is Subjected to an extraction process as depicted in FIG. 1. In accordance with this embodiment, 0209. In an alternate embodiment of the invention, plant three basic extraction processes are performed in sequence material is extracted with an alcoholic solvent, such as ethanol, methanol. 1-propanol, 1-butanol, 2-propanol, 2-bu to generate potential extracts A, B and C. tanol, 2-methyl-1-propanol, 2-methyl-2-propanol, glycerine, 0204. In other embodiments of the present invention, ethylene glycol, propylene glycol, diethylene glycol, dipro greater or fewer extraction processes are contemplated. For pylene glycol or 1,3-butylene glycol or a combination of example, in an alternative embodiment, the plant material is alcoholic solvents. In one embodiment, a combination of subjected to an extraction process as depicted in FIG. 5. In ethanol and methanol is used as the alcoholic Solvent, accordance with this embodiment, the plant material is, wherein the range of ethanol:methanol is between about Subjected to two separate extraction processes concurrently 50:50 and about 85:15. In another embodiment, a glycol is resulting in two separate potential extract AS. used as the alcoholic solvent. In a further embodiment, the 0205 Regardless of the number of extraction processes, plant material is contacted with an alcoholic solvent for a the procedure for each extraction process entails contacting time period between about 10 minutes to one hour at a the solid plant material with a solvent with adequate mixing temperature between about 4 to about 25°C. and for a period of time Sufficient to ensure adequate 0210. In an alternate embodiment, plant material is exposure of the solid plant material to the solvent such that extracted with an alcoholic solvent in combination with a inhibitory activity present in the plant material can be taken co-solvent, which may be aqueous or organic. In one up by the solvent. Typically, the extraction procedures are embodiment, a combination of ethanol and water is used as conducted over a period of time between about 10 minutes the solvent, wherein the range of ethanol: water is between and about 24 hours at a temperature between about 4°C. and about 50:50 and about 85:15. In another embodiment, a about 50° C. Other times and temperatures may be employed combination of a glycol and water is used as the Solvent, in the extraction process as described above. Adequate wherein the range of glycol: water is between about 95:5 and contact of the solvent with the plant material can be encour about 50:50. aged by shaking the Suspension. The liquid fraction is then separated from the solid (insoluble) matter resulting in the 0211. In an alternate embodiment, plant material is generation of two fractions: a liquid fraction, which is a extracted with an organic solvent. Such as diethylether, potential extract, and a Solid fraction. Separation of the hexane, heptane, dichloromethane, or ethylacetate. In one liquid and solid fractions can be achieved by one or more embodiment, dichloromethane is used as the solvent and the standard processes known to those skilled in the art. plant material is shaken for one to twenty-four hours with the solvent. 0206. In accordance with the embodiment depicted in FIG. 1, the extraction process is then repeated with a second 0212. Once the potential extracts have been isolated, they and a third solvent. Solvents A, B and C in FIG. 1 generally can be tested directly (after being dissolved or dispersed in represent separate classes of solvents, for example, aqueous, a suitable solvent) for their ability to inhibit skin EP activity, alcoholic and F organic. The solvents can be applied in or they may be subjected to further procedures as described specific order, for example, a polar to non-polar order or in below and outlined in FIGS. 2 and 6. For example, the a non-polar to polar order. Alternatively, the solvents can be potential extracts can be subjected to procedures to remove applied in a random sequence. In all cases, however, the fatty acids or chlorophyll components that may interfere solid matter should be dried prior to contact with the with the protease activity or other assays. Various proce Subsequent solvent. dures known in the art may be employed. In one embodi ment, one or more additional partitioning step using an 0207. The plant material employed in the extraction organic solvent, Such as hexane, heptane or ethyl acetate, is process can be the entire potential plant, or it can be one or included. The liquid potential extract can be concentrated more distinct tissues from a plant, for example, leaves, and Solubilised in an appropriate solvent prior to the one or seeds, roots, stems, flowers, and the like, or various com more partitioning step, if desired. binations thereof. The plant material can be fresh, dried or frozen. If desired, the plant material can be treated prior to 0213 The present invention contemplates that the extrac the extraction process in order to facilitate the extraction tion process may be carried out on various scales including process. Typically such treatment results in the plant mate known large, medium and Small-scale methods of preparing rial being fragmented by some means such that a greater eXtractS. US 2007/01 22492 A1 May 31, 2007 24

0214) Determination of Skin Extracellular Protease gamma linolenic acid (6.9,12-Octadecatrienoic acid, Sigma Inhibiting Activity L-2378) (stress G) or arachidonic acid-(5,8,11,14-Eicosatet 0215. Following the extraction process, the potential raenoic acid, Sigma A-3925) (stress A) (400 uM in water extracts are tested for their ability to inhibit one or more skin with 0.125% (v/v) Triton X-100) to completely cover the EPs selected from the group of MMP-1, MMP-2, MMP-3, leaves. Twenty to twenty-four hours after the stress, plants MMP-9 and HLE, using one of a variety of techniques were harvested. known in the art including, but not limited to, those 0223 Harvest Solid S1 and Optional Storage Treatment: described herein. Those plant extracts that decrease the More than 4 grams of leaves, stems, fruit, flowers, seeds or activity of at least one skin EP by at least 20% are selected other plant parts were harvested from stressed or non for further testing. In one embodiment of the present inven stressed plants and frozen immediately in dry ice, then tion, plant extracts that inhibit the activity of one or more of transferred as soon as possible to a -20°C. freezer until use. MMP-1, MMP-2, MMP-3, MMP-9 and HLE by at least 30% Plant materials may be stored at -20° C. for than a year are selected. In another embodiment, plant extracts that without losing inhibitory activity. Temperature was moni inhibit the activity of one or more of MMP-1, MMP-2, tored to ensure a constant condition. MMP-3, MMP-9 and HLE by at least 40% are selected. In another embodiment, plant extracts that inhibit the activity 0224) Stressed and non-stressed plant specimens were of one or more of MMP-1, MMP-2, MMP-3, MMP-9 and collected as wet samples and stored at -20°C. for various HLE by at least 50% are selected. periods of time, and were submitted to a process which generates 3 subfractions: aqueous, ethanolic and organic 0216) In order to determine whether the potential extracts fractions. The complete extraction process was performed in inhibit a skin EP, the extracts can be tested against an a continuous cycle using the following steps. An initial 5 g individual skin EP or against a panel comprising two or of plant specimen was homogenized in liquid nitrogen with more of MMP-1, MMP-2, MMP-3, MMP-9 and HLE. a blender. The resulting powder was weighed. Similarly, the extracts can be tested individually or a plu rality of extracts can be tested simultaneously using high 0225 Extraction Process I Aqueous Extraction: To throughput assays, as known in the art. Simultaneous testing each 4.5 grams of plant powder, 12 ml of a cold solution of of a plurality of extracts maximizes the number of extracts 100 mM Tris, pH 7.0 was added. The mixture was thor that can be tested in a set period of time and thus decreases oughly vortexed for 2 minutes. The mixture was kept on ice the overall time for the screening process. for 30 minutes and vortexed after each 10 minute period of time. The sample was centrifuged in a CorexTM 30 ml tube 0217 Cellular Screening of Extracts for 5 minutes at 4500 rpm. The resulting supernatant was 0218. Those extracts identified as being capable of inhib decanted in a 15 ml tube after filtration with a MiraclothTM iting one or more of MMP-1, MMP-2, MMP-3, MMP-9 and filter. This extract represents Potential Extract A in FIG. 1. HLE are subsequently screened for their ability to affect one The pellet, referred to as Solid S2, was kept for ethanolic or more cellular activities in skin cells. Such cellular activi extraction. ties include, for example, attenuating the breakdown of a 0226. The aqueous extract (Potential Extract A) was structural component of the ECM (i.e. collagen, fibronectin, further purified in order to determine its EP inhibition fibrillin and/or elastin); attenuating endothelial cell migra capability. The Potential Extract A was purified by size tion; increasing collagen production; attenuating UV-in exclusion chromatography, wherein the aqueous extract was duced extracellular protease activity and/or attenuating trac chromatographed on a calibrated Sephadex G-25 column tional forces generated by fibroblasts. The extracts can be (1x10 cm) using a 20 mM Tris-HCl, 150 mM. NaCl, pH 7.5 tested using standard methods such as those described buffer as eluant. Fractions corresponding to compounds that above. appeared to have a molecular weight (NW) less than 1500 0219. Further Testing daltons (D) were pooled to constitute the purified aqueous 0220. The extracts identified by the above process may be eXtract. Submitted to other standard tests, such as cytotoxicity tests, 0227 Prior to analysis of the aqueous extract for inhibi stability tests, bioavailability tests and the like, to determine tory activity as described in Example II, the extract was their suitability for inclusion in a dermatological formulation treated with 10% gelatine-Sepharose (Pharmacia Biotech, of the invention. Exemplary tests are described above. Uppsala, Sw.) in order to remove unspecific enzyme ligands. 0221) To gain a better understanding of the invention To 1 mL of extract, 100 uL of gelatine-Sepharose resin was described herein, the following examples are set forth. It added in a microassay tube, the solution in the tube was should be understood that these examples are for illustrative mixed, kept on ice for 30 minutes, and then centrifuged 5 purposes only. Therefore, they should not limit the scope of minutes at 5,000 rpm. The supernatant was removed and this invention in any way. used directly for assays. 0228. Extraction Process II—Alcoholic Extraction: To EXAMPLES the pellet, Solid S2, collected from the previous aqueous extraction, 12 ml of cold ethanol:methanol (85:15) was Example I added and the mixture was thoroughly vortexed for 2 minutes. The mixture was kept on ice for 30 minutes and Preparation of Stressed and Non-stressed Plant vortexed every 10 minutes. The sample was centrifuged in Extracts (Method A) a Core XTM 30 ml tube for 5 minutes at 4,500 rpm. The 0222 Optional Pre-Harvest Treatment: Aerial parts of a resulting Supernatant was decanted in a 15 ml tube after living plant were sprayed with an aqueous solution of filtration with a MiraclothTM filter. The pellet, referred to as US 2007/01 22492 A1 May 31, 2007

Solid S3, was kept for the Subsequent organic extraction. MMP-1, HT-1080 (ATCC, Manassas, Va.) for MMP-2 and This extract represents Potential Extract B. THP-1 (ATCC, Manassas, Va.) for MMP-9) as described in literature and based on protocols found in I. M. Clark: 0229. The ethanolic extract, Potential Extract B, was <>, Humana Press purified by liquid/liquid extraction prior to analysis by (2001). Recombinant human MMP-3 was overexpressed in enzymatic assay. For this purpose, 1 ml of ethanolic extract E. coli and purified according to Windsor L. J. Steele DL was evaporated under vacuum, dissolved in 150 ul of (2001), Methods Mol Biol 151:191-205. Proteolytic activity dimethylsulfoxide (DMSO), and completed to a final vol of these proteases was evaluated with the assay based on the ume of 1.5 ml with Tris buffer (final concentration: Tris-HCl cleavage of auto-quenched peptide substrate: (MCA-Pro mM; pH 7.5). Four ml of hexane was added to the Tris phase Leu-Gly-Leu-Dpa-Ala-Arg-NH TFA Dpa=N-3-(2,4-dini in a glass tube and the tube was thoroughly vortexed, then trophenyl)-L-2,3-diaminopropionyl) for MMP-1, -2, and allowed to form a biphasic liquid. The organic phase was -9; and, MCA-Arg-Pro-Lys-Pro-Val-Glu-Nva-Trp-Arg-Lys removed and the extract was submitted to a second round of (DNP). NH (DNP=2,4-dinitrophenyl; Nva=L-norvaline) liquid/liquid extraction. The aqueous phase was removed for MMP-3 (Calbiochem, San Diego, Calif.). In the intact and treated with 10% gelatine-Sepharose (Pharmacia Bio peptide. Dpa or DNP quenches the MCA fluorescence. tech, Uppsala, Sw) to remove non-specific enzyme ligands Cleavage of the peptide causes release of the fluorescent prior to conducting Subsequent assays. To 1 ml of extract, MCA group which was then quantitated on a fluorometer 100 uL of gelatine-Sepharose resin was added in a microas (Gemini XS, Molecular Devices, Sunnyvale, Calif.). The say tube, the tube was mixed, kept on ice for 30 minutes, and assay was performed in TNCZ assay buffer (20 mM Tris then centrifuged 5 minutes at 5,000 rpm. Supernatant was HCl; NaCl 150 mM; CaCL 5 mM, ZnCl, 0.5 mM; pH 7.5) removed and used directly for assays as described in with human purified proteases (I. M. Clark: Matrix metal Example II. loproteinases protocols, Humana Press (2001)). The sub 0230. Extraction Process III Organic Extraction: To the strate, primarily dissolved in DMSO was then redissolved in pellet, Solid S3, collected from the previous ethanolic TNCZ buffer for the assay. In a typical assay, 10 ul of extraction, 12 ml of cold dichloromethane was added and the purified enzyme (1-50 ng) and 5 ul of dissolved substrate mixture was thoroughly vortexed for 2 minutes. The mixture (final concentration of 10 uM) was mixed in a final volume was kept on ice for 30 minutes and vortexed after each 10 of 75ul (completed with TNCZ). All assays were performed minutes period. The sample was centrifuged in a CorexTM 30 in 96 well plate and the reaction was started by the addition ml tube for 5 minutes at 4,500 rpm. The resulting superna of Substrate. Assays were measured (excitation 325 nm, tant was decanted in a 15 ml glass tube after filtration with emission 392 nm) for 20, 40 and 60 minutes. a MiraclothTM filter. The final pellet was discarded. The organic solvent was evaporated under vacuum and the phase 0235 Measurement of Human MMP-9 or Human Leu was dissolved with dimethylsulfoxide (DMSO). This extract kocyte Elastase (HLE) Activity. Using the FASC Assay represents Potential Extract C, which was further purified by 0236 Human leukocyte elastase was obtained from Cal Solid phase extraction prior to analysis by enzymatic assay. biochem (San Diego, Calif.). Human MMP-9 was purified 0231. In order to assay the Potential Extract C, the as previously described. The assay was based on the method organic extract was diluted 1:10 in a solution of described in Canadian Patent No. 2,189,486 (1996) and by DMSO:Methanol:Tris (20 mM, pH 7.5) (10:50:40) (Solu St-Pierre et al., (Cytometry (1996) 25:374-380. For the tion A), i.e., 220 ul of extract was added to 2.0 ml of solution assay, 5 ul of the purified enzyme (1-100 ng), 5 ul of A. After 10 seconds of vigorous vortex, the mix was concentrated buffer solution (20 mM Tris-HCl, NaCl 150 sonicated for 10 seconds. Dissolved extracts were subse mM: CaCL 5 mM; ZnCl, 0.5 mM; pH 7.5), and 5 ul of quently applied to a solid phase extraction plate (Discovery gelatine-FITC beads were typically used in a final volume of SPE-96, Sigma Chemical Co., St-Louis, Mo.). After initial 100 ul. The assay was performed by incubation of the conditioning of the columns with 1 ml of methanol, columns reaction mixture for 90 minutes at 37° C. The reaction was were equilibrated with Solution A, and extract samples were stopped by the transfer of the mix in 0.5 ml of 20 mM Tris, deposited on the columns. Elution was completed with 150 mM. NaCl; pH 9.5 buffer. This tube was analyzed in a solution A (final volume of 2 ml) and this fraction was used flow cytometer (Epics MCL. Beckman Coulter, Missis directly in assays as described in Example II. sauga, Ontario) as described in Canadian Patent No. 2,189, 486. Example II 0237 Measurement of HLE Activity with a Fluorogenic Proteic Substrate In vitro Enzyme Inhibition Assays 0238 HLE was obtained from Calbiochem (San Diego, 0232 The inhibitory activity of sample compositions Calif.). The activity of HLE was measured by an assay based towards human MMP-1, human MMP-2, human MMP-3, on the increase of fluorescence of a proteic substrate (beta human MMP-9 and/or human leukocyte elastase (HLE) casein) heavily labelled with Alexa-488 dye (Molecular were determined using either fluorogenic Substrates or the Probes, Eugene, Or). The substrate, when highly labelled FASC assay. with the dye, will almost quench the dye fluorescence. Cleavage of the substrate will result in an increase of the 0233. Measurement of Human MMP-1, -2, -3 and -9 fluorescence which can be measured with a spectrofluorom Activity with Fluorogenic Peptidic Substrates eter, and which was proportional to protease activity. Typi 0234 MMP-1, -2, -9 were purified from natural sources cally, 10 ul of purified HLE (10-50 ng) and 10 uL of (human immortalized cell lines: 8505C (Deutsche Sam beta-casein-Alexa488 (100 ng) were assayed in final volume mlung von Mikroorganismen und Zellkulturen GmbH) for of 75 ul adjusted with 20 mM TNCZ buffer. The reaction US 2007/01 22492 A1 May 31, 2007 26 was performed as already described except that the fluores cence was read at excitation 488 nmfemission 525 nm. TABLE 1-continued wavelengths. Inhibition of MMP-1 by Plant Extracts 0239). Inhibition Assay for Plant Extracts Inhibition 0240 Before a typical assay, aqueous extracts prepared Latin Name Stress Extract (%) as described in Example I were preincubated with 1:10 of A. icul A. S 30.4 gelatine-Sepharose 4BTM for 30 minutes to remove fluores- t; C'C' iii. A. 4 36.4 cence quenching. For the ethanolic extract, an initial hexane Allium cepa A. O 61.4 extraction was performed and samples were treated with Allium grande A. R 46.5 1:10 of gelatine-Sepharose 4BTM to remove quenching. then porrain A 5. it in porrain 0241. In a typical fluorescent assay, 10 ul of purified Allium sativum A. O 42.5 enzyme at concentrations previously mentioned for the then t A s enzymatic assay, 5 ul of dissolved fluorogenic peptide or 10 Aiumit in SchoenoprastinTiberosum A. R 29.9 ul of dissolved fluorescent proteic substrate (final concen- Aium Tiberosum A. O 1OO.O tration of 10 uM) and 40 uL of the aqueous, ethanolic or Althaea officinalis A. S 21.6 organic extract to be tested were mixed in a final Volume of AngelicaAnthemis archangelicanobiis A. SR 45.934.5 75 ul (completed with TNCZ for fluorogenic peptide sub- Aralia nudicatiis A. O 1OO.O strate assay or 20 mM citrate pH 3.3 buffer for fluorescent Armoracia rusticana A. O 31.2 protein substrate assay). All assays were performed in 96 Armoracia rusticana A. S 39.7 well plate and the reaction was started by the addition of tput melanocarpa Sier sp substrate. Assays were measured (excitation 325 nm, emis- Beckmannia eruciformis A. O 24.1 sion 392 nm for peptide and excitation 488 mm/emission 525 Beta vulgaris A. R 41.2 nm wavelengths for protein) for 20, 40 and 60 minutes. Beta vulgaris spp. Maritima A. O 44.1 Activity and inhibition values were determined from the Brassica napus A. O 26.3 in fl Brassica oleracea A. S 28.6 1CaS 1 OSCC Brassica oleracea A. R 33.8 0242 For the FASC assay, 35 ul of the treated extract E.Brassica alsool A. OR 1OO.O61.4 prepared as described in Example I, 5 Jul of the purified Calamintha nepeta A. R 40.2 enzyme prepared as described previously, 5 Jul of concen- Cameia sinensis A. O 39.3 trated buffer solution (TNCZ), and 5 ul of gelatine-FITC capsicum annuum A. R 34.3 beads were typically used. The initial step of the assay was capsicumCapsicum annuitinfinitesCens A. OR 39.488.3 the incubation of the reaction without beads for a 30 minutes Chenopodium bonus-henricus A. O 1OO.O period on ice to allow the binding of inhibitors to enzyme. Chenopodium bonus-henricus A. R 37.3 Fluorescent beads were added and the reaction mix was C quinoa A g incubated for 90 minutes at 37°C. The reaction was stopped Ciinternatin A. R 22.0 by transfer of the mix in 0.5 ml of 20 mM Tris, 150 mM Cichorium intybus A. S 66.9 NaCl; pH 9.5 buffer. This tube was analyzed in the flow Citritius ianatus A. O 41.9 cytometer (Epics MCL. Beckman Coulter, Mississauga, Cornus Canadensis A. S 73.O Crataegus sp A. O 1OO.O Ontario) as described in Canadian Patent Application No. Clictimis Anguria A. S 34.2 2,189,486 (1996). Cucurbita moschata A. O 27.3 0243 Results of the inhibition------studies are shown in CymbopognCucurbita pepo citratus A. O 1OO.O84.9 Tables 1-5 for aqueous (O), ethanolic (R) and organic (S) Cymbopogon citratus A. R 22.1 extracts from exemplary stressed (A: Arachidonic acid and Cypertis esculenius A. R 25.8 G: Gamma-linolenic acid) and non-stressed (T) plant CypertisDactylis glomerataesculenius A. O 28.125.5 sources. The inhibition is reported aS percentage (%) of Daiiciis Caroia A. O 43.4 inhibition of Substrate degradation as compared with Sub- Daiiciis Caroia A. R 1OO.O strate degradation in the absence of the extract. Percentage pite A 3. inhibition was calculated according to the formula: Ericairca palustrisvesicaria A. R 33.7 Percentage (%) inhibition=EA-EEEAx100 Eschscholzia californica A. O 61.1 Eschscholzia californica A. R 74.1 0244 wherein E is the protease activity in the absence of Filipendula rubra A. O 51.7 the plant extract and E is the protease activity in the foeniculum vulgare A g s: ragaria X ananassa presence of the extract. Fragaria Xananassa A. S 40.6 Fragariax ananassa A. R 28.3 TABLE 1. Gainsoga ciliata A. R 29.7 Gaium odoratum A. 6 48.8 Inhibition of MMP-1 by Plant Extracts Gaultheria hispidula A. R 23.9 Glycine max A. R 24.7 Inhibition Glycine max A. S 29.6 Latin Name Stress Extract (%) Glycine max A. O 1OO.O Guizotia abyssinica A. S 39.4 Achillea millefolium A. O 22.2 Hamamelis virginiana A. R 49.1 Acorus calamus A. O 1OO.O Heianthus Tuberosus A. O 95.9 Actinidia arguita A. O 56.4 Heliotropium arborescens A. R 2S.O US 2007/01 22492 A1 May 31, 2007 27

TABLE 1-continued TABLE 1-continued Inhibition of MMP-1 by Plant Extracts Inhibition of MMP-1 by Plant Extracts Inhibition Inhibition Latin Name Stress Extract (%) Latin Name Stress Extract (%) Hordeum hexastichon A. O 1OO.O Trifolium pannonicum A. R 27.7 Hordeum vulgare A. O 46.2 Trifolium repens A. R 79.5 Hordeum vulgare subsp. Vulgare A. O 43.8 Vaccini in augustifolium A. R 52.5 Initia heienium A. O 25.8 Vaccinum macrocarpon A. O 64.5 Lathyrus Saivais A. O 27.1 Vicia Saiva A. O 60.8 Leonirits cardiaca A. O 34.4 Vicia Saiva A. R 28.6 Levisticum officinale A. R 31.7 Vicia viliosa A. R 64.7 Lolium multiflorum A. O 39.0 Vicia viliosa A. O 57.3 Lotus corniculatus A. O 1OO.O Vigna Sesquipedalis A. O 33.0 Malva Sylvestris A. R 22.8 Vigna Sesquipedalis A. R 24.4 Matricaria rectitita A. O 25.1 Vignatinguiculata A. R 20.6 Matteucia pensylvanica A. R 48.1 Vitia spp A. R 72.6 Medicago Saiiva A. R 25.1 Vitia spp A. O 1OO.O Melissa officinalis A. O 1OO.O Zea Mays A. R 99.2 Mentha piperita A. O 60.1 Zea Mays A. O 1OO.O Mentha suaveolens A. O 35.1 Abelnochtis escientiis G R 37.6 Nepeta cataria A. O 1OO.O Aconium napeius G O 1OO.O Nicotiana rustica A. R 20.7 Allium ampeioprastin G R 33.4 Origantim vulgare A. R 6O.S Allium ascalonictim G R 31.5 Origantim vulgare A. O 73.2 Allium cepa G O 34.4 Perilla frutescens A. R 74.4 Allium cepa G R 36.4 Perilla frutescens A. O 92.4 AAium sativum G R 53.2 Petroselinum crispum A. R 77.4 Aium Tiberosum G R 68.3 Phacelia tanacetifolia A. R 52.8 Althaea officiana is G O 47.7 Phaseolus coccineus A. R 20.9 Althaea officinalis G S 30.7 Phaseolus coccineus A. S 34.2 Althaea officinalis G S 44.3 Phaseolus Vulgaris A. S 29.2 Althea officinalis G R 83.6 Phaseolus vulgaris A. R 56.1 Anethium graveolens G S 44.3 Phaseolus Vulgaris A. R 60.0 Apium graveolens G R 27.7 Phaseolus Vulgaris A. O 1OO.O Armoracia rusticana G O 51.8 Phlox paniculata A. O 1OO.O Armoracia rusticana G S 47.1 Pimpinella anisum A. S 1OO.O Aronia melanocarpa G S 66.5 Pimpinella anisum A. R 72.2 Artemisia dractinctius G S 79.0 Plantago Coronopus A. R 23.7 Artemisia dractinctius G R S.O.3 Plectranthus sp. A. O 2S.O Asparagus officinalis G O 96.4 Poa compressa A. O 31.5 Bellis perennis G R 44.1 Potentiiia anserina A. R 71.2 Beta vulgaris spp. Maritima G R 43.7 Pysalis ixocarpa A. R 32.1 Beta vulgaris spp. Maritima G O 34.9 Raphantis raphanistrain A. O 31.5 Betula glandulosa G S 40.8 Raphantis Saivais A. O OO.O Borago officinalis G O 30.3 Raphantis Saivais A. O 30.2 Borago officinalis G R 29.7 Rheum officinale A. O 79.1 Brassica cepicepa G R 21.9 Rheum rhabarbarium A. R 22.9 Brassica oleracea G O 33.6 Rheum rhabarbarium A. R 32.8 Brassica oleracea G O 1OO.O Ribes nigrum A. O OO.O Brassica rapa G O 42.5 Ribes nigrum A. R OO.O Brassica rapa G R 40.2 Ribes Saivum A. R 48.6 Calamintha nepeta G O 28.7 Ribes Sylvestre A. S 26.5 Calendula oficinalis L. G O 1OO.O Ribes tiva-crispa A. R OO.O Cameia sinensis G O 46.4 Rubus Canadensis A. R 46.1 Campaniiia raptinctiitis G R 27.2 Rubus Canadensis A. R 53.1 Capsella bursa-pastoris G R 24.1 Rubus idaeus A. R OO.O Capsicum annum G O 36.0 Salvia officianalis A. O OO.O Chaerophyllum bulbosum G R 38.9 Saivia Sciarea A. S 43.8 Chenopodium quinoa G O 1OO.O Satureia montana A. R OO.O Cichorium intybus G S 44.6 Soianum duticamara A. S 43.8 Circium arvense G R 30.3 Soianum melanocerastin A. R 37.2 Citritius ianatus G R 21.2 Soianum tuberosum A. R OO.O Cucurbita pepo G O 59.5 Sorghum dochna A. O OO.O Cucurbita Pepo G O 40.2 Stachys byzantina A. S 28.9 Cuminum cyminum G R 25.5 Steiaria media A. S 33.1 Cymbopogon citratus G R 33.7 Tanacetain parthenium A. O 28.9 Daitira Stranonium G O 73.5 Tanacetum vulgare A. R 76.O Daiiciis Caroia G O 86.0 Taraxacum officinale A. O 65.7 Daiiciis Caroia G O 27.9 Thymus praecox. Subsp. arcticus A. O 642 Dryopteris filix-mas G O 21.9 Thymus praecox. Subsp. arcticus A. R 88.2 Erysimum perofskianum G O 24.4 Thymus vulgaris A. R 42.7 Fagopyrim esculenium G O 1OO.O Thymus X citriiodorus A. O 34.7 Foeniculum vulgare G O 28.0 Trichosanthes kirilowii A. R 31.8 Foeniculum vulgare G R 57.3 Trifolium hybridum A. R 96.O Gaultheria hispidula G O 44.2 Trifolium incarnatum A. R 1OO.O Gaultheria procumbens G R 94.8 US 2007/01 22492 A1 May 31, 2007 28

TABLE 1-continued TABLE 1-continued Inhibition of MMP-1 by Plant Extracts Inhibition of MMP-1 by Plant Extracts Inhibition Inhibition Latin Name Stress Extract (%) Latin Name Stress Extract (%) Giechona hederacea G O 25.5 Salvia officinalis G R 1OO.O Glycine max G S 1OO.O Santoina G R 28.1 Glycyrrhiza glabra G O 24.9 Satureia hortensis G R 1OO.O Guizotia abyssinica G R 30.3 Satureia repandra G O 57.1 Helenium hoopesii G O 28.6 Scrophtiaria nodosa G R 41.6 Helianthus annuit is G O 33.6 Scutelaria laterifiora G S 72.1 Heianthus tuberosus G O 54.4 Siim Sisartin G O 99.7 Hordeum vulgare G O 28.8 Soianum duticamara G R 65.4 Hordeum vulgare subsp. Vulgare G R 28.1 Soianum melanocerastin G R 32.4 Hypericum henryi G R 80.0 Soianum melogena G O 1OO.O Iberis amara G O 44.6 Soianum tuberosum G S 46.4 Lactica Saiiva G R 25.3 Sorghum cafiorum G R 1OO.O Lathyrus Sylvestris G O 90.2 Sorghum dochna G R 51.4 Lavandhiia angustifolia G R 22.5 Sorghum dochna G R 39.6 Lepidium Sativum G S 29.5 Sorghum Sudanense G O 97.4 Levisticum officinale G O 1OO.O Stachys byzantina G O 41.4 Lolium multiflorum G O 24.9 Steiaria media G O 33.8 Lolium multiflorum G R 27.1 Symphytum officinale G O S2.O Lotus corniculatus G O 52.2 Tanacetain parthenium G O 79.1 Lycopersicon esculenium G R 24.4 Tanacetum vulgare G O 1OO.O Lycopersicon pinpineliifolium G R 30.3 Taraxacum officinale G S 25.9 Maius hupehensis G R 65.8 Teucrium chamaedrys G O 1OO.O Maiva verticiliata G R 43.1 Teucrium chamaedrys G R 48.0 Matricaria rectitita G S 1OO.O Thymus praecox. Subsp. arcticus G R 73.1 Matteucia pensylvanica G R 57.5 Thymus X citriiodorus G O 52.2 Melissa officinalis G O 28.5 Trichosanthes kirilowii G O 35.9 Mentha piperita G O 36.0 Trifolium hybridum G R 76.O Mentha spicata G S 20.3 Trifolium incarnatum G R 73.4 Mentha spicaia G S 26.O Trifolium pannonicum G R 24.8 Mentha suaveolens G O 6O.S Trifolium repens G R 48.5 Nepeta cataria G O 24.1 Triticosecale spp. G R 48.5 Nicotiana rustica G R 28.1 Triticum spelta G R 22.9 Nicotiana tabacum G R 40.6 Tropaeoluim mails G S 23.4 Oenothera biennis G R 28.4 Urtica dioica G O 96.4 Oenothera biennis G O 1OO.O Vaccinium corymbosum G S 60.7 Origantim vulgare G S 1OO.O Vaccinium corymbosum G R 61.4 Origantim vulgare G O 20.1 Vaccini in anguistifolium G R 54.7 Origantim vulgare G O 85.4 Vicia Saiva G R 68.8 Oryza Sativa G R 53.3 Vicia Saiva G O 31.5 Panax quinquefoils G S 1OO.O Vicia viliosa G O 1OO.O Panicum miliaceum G S 1OO.O Vicia viliosa G R 35.5 Passiflora caerula G O 20.9 Vigna Sesquipedalis G R 23.0 Pastinaca Saiiva G R 68.4 Vitia spp G R 36.9 Pastinaca Saiiva G O 1OO.O Withania somnifera G O 44.0 Pennisetum alopectiroides G R 1OO.O Xanthium strumarium G R 37.6 Petroselinum crispum G R 73.O Zea mayS G O 1OO.O Phaiaris canariensis G O 1OO.O Aconium napeius R 1OO.O Phaseolus coccineus G R 29.9 Agarict is bisports R S8.9 Phaseolus coccineus G R 67.6 Agarict is bisports O 1OO.O Phaseolus coccineus G O 32.4 Allium ampeioprastin R 43.3 Phaseolus vulgaris G R 33.4 Allium ascalonictim R 34.5 Phaseolus vulgaris G R 60.2 Allium cepa R 53.5 Phaseolus vulgaris G R 22.3 Allium cepa O 45.8 Phaseolus vulgaris G O 87.7 Allium grande R 43.2 Phlox paniculata G O 89.3 Allium Schoenoprastin R 47.1 Physalis pruinosa G O 37.0 Aium Tiberosum R 74.6 Plantago Coronopus G R 48.1 Aium Tiberosum O 33.6 Plantago major G O 47.0 Aloe Vera R 34.1 Plectranthus sp. G O 97.2 Althaea officinalis S 47.8 Potentiiia anserina G R 22.O Amelianchiera initoia R 59.1 Prunella vulgaris G O 21.2 Ananas Conostis O 1OO.O Raphantis Raphanistrain G O 95.9 Anthemis nobiis O 22.7 Raphantis Saivais G O 67.7 Anthriscus cerefolium O 56.8 Reseda Odorata G O 40.6 Apium graveolens R 29.8 Rheum officinale G O 82.1 Aralia nudicatiis O 1OO.O Rheum rhabarbarium G R 48.1 Armoracia rusticana O S8.9 Ribes Nigrum G R 1OO.O Artemisia dractinctius O 1OO.O Ribes Sylvestre G O 42.9 Asparagus officinalis R 25.2 Ricinits communis G O 73.5 Atriplex hortensis R 44.7 Rubus Phoenicaiasius G R 31.4 Bellis perennis R S8.1 Rita graveolens G R 1OO.O Beta vulgaris R 37.3 US 2007/01 22492 A1 May 31, 2007 29

TABLE 1-continued TABLE 1-continued Inhibition of MMP-1 by Plant Extracts Inhibition of MMP-1 by Plant Extracts Inhibition Inhibition Latin Name Extract (%) Latin Name Extract (%) Betula glanditiosa 23.5 Melissa officinalis Boletus eduis 642 Melissa officinalis Brassica iuncea 35.6 Mentha suaveoiens 39.7 Brassica naptis 1OO.O Nigella Saiiva S8.9 Brassica oleracea 33.2 Nigella Saiiva 1OO.O Brassica oleracea 49.7 Ocimum Basilicum 1OO.O Cameia sinensis 24.7 Origantin majorana 41.5 Cameia sinensis 45.7 Origantim vulgare 29.8 Canna editiis 26.2 Origantim vulgare 33.1 Cartin carvi 1OO.O Panax quinquefoils 75.2 Chaerophyllum bulbosum 40.9 Passifiora spp. 32.O Chrysanthemun coronarium (Chip Suey) 48.1 Pastinaca Saiiva 20.8 Chrysanthemum coronarium 29.9 Perroselinum crispum SS.4 Chrysanthemum coronarium 1OO.O Petroselinum crispum 76.1 Cichorium endivia 2O.S Petroselinum crispum 24.1 Cichorium endivia 21.9 Peucedanum or'easelinum 21.O Cichorium intybus SO.6 Phacelia tanacetifolia 48.6 Cichorium intybus 31.7 Phaiaris canariensis 56.4 Cichorium intybus S2.9 Phaseolus coccineus 22.7 Citritius ianatus 1OO.O Phaseolus mingo 47.4 Citrus paradisi 40.6 Phaseolus vulgaris 40.O Cocos nucifera 27.2 Phaseolus vulgaris 29.4 Cornus Canadensis 44.9 Phoenix dactylifera 46.3 Crithmin maritimum 32.3 Physalis ixocarpa goldie out poirpre 28.9 Cuctimis anguria 22.6 Phytolacca americana OO.O Cucurbita moschata 33.5 Plectranthus sp. 73.8 Cucurbita moschata (Early Butternut) 32.3 Pleurotus spp. OO.O Cucurbita pepo 89.0 Poa compressa 22.3 Cuminum cyminum S4.3 Poa pratensis 73.1 Curcuna Zedoaria 1OO.O Populus Tremula OO.O Cymbopogon citratus 42.6 Prunella vulgaris 38.0 Datura metel 24.8 Psoralea corylifolia 96.4 Datura metel 25.5 Pteridium aquilinum OO.O Dioscorea batatas 1OO.O Raphantis raphanistrain OO.O Dipsacus sativus 85.0 Raphantis Saivais 33.7 Dryopteris filix-mas 46.4 Raphantis Saivais 28.0 Erigeron Canadensis 1OO.O Raphantis Saivais OO.O Erica vesicaria 30.9 Reseda luteola 69.6 Erysimum perofskianum 23.0 Reseda Odorata 51.8 Eschscholzia californica 37.8 Rheum officinale 46.7 Eschscholzia californica 20.8 Rheum officinale OO.O Fagopyrim esculenium 1OO.O Ribes nigrum 3O.O Fagopyri in tariarictim 78.5 Ribes Saiivum 61.7 Foeniculum vulgare 63.4 Ribes Sylvestre 75.4 Foeniculum vulgare 27.2 Ricinits communis OO.O Forsythia X intermedia 32.O Rosmarinus officinalis 29.0 Fragaria X ananassa 33.0 Rubus Canadensis 86.1 Gainsoga ciliata 25.8 Sabai Serrutiaia OO.O Gaultheria procumbens 46.8 Salvia officinalis OO.O Hedeona pullegioides 73.6 Sambucus Canadensis 24.8 Heianthus tuberosus 39.3 Satureia montana OO.O Hordeum vulgare 32.4 Satureia repandra 27.2 Huntilus lupuits 21.1 Satureia repandra 36.4 Hypericum henryi 29.3 Satureia repandra 42.O Hyperictim perforatin 42.7 Scrophtiaria nodosa 68.8 Iberis amara 29.5 Secale cereale 1OO.O Ipomea aquatica 22.9 Setaria italica 23.2 Lathyrus Saivais 69.4 Silybum marianum 73.5 Latiri is nobilis 70.2 Soianum melongena 20.1 Lavandhiia iaiifolia 1OO.O Soianum tuberosum 24.4 Lens culinaris Subsp. Culinaris 70.2 Solidago virgau rea 71.4 Lepidium sativum 1OO.O Sorghum dochna 22.5 Levisticum officinale 1OO.O Stachys byzantina 39.2 Lolium multiflorum 35.1 Steiaria media 43.3 Linaria annia 1OO.O Symphytum officinale Lycopersicon pinpineliifolium 24.4 Tanacetain parthenium Maius hupehensis 73.1 Tanacetum vulgare 32.5 Maius sp. 80.9 Taraxacum officinale 27.8 Malva Sylvestris 34.7 Teucrium chamaedrys Malva Sylvestris 1OO.O Teucrium chamaedrys Manihot escuienia 33.0 Thalipsi arvense 21.2 US 2007/01 22492 A1 May 31, 2007 30

TABLE 1-continued TABLE 2-continued Inhibition of MMP-1 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin Name Stress Extract (%) Latin name Stress Extract (%) Thymus praecox. Subsp. arcticus R 60.9 Brassica naptis A. O 28.5 Tragopogon porrifolium R 24.6 Brassica naptis A. S 52.4 Trifolium incarnatum R 33.7 Brassica naptis A. R 82.4 Trifolium pannonicum R 72.4 Brassica nigra A. O 29.2 Trifolium repens R 72.4 Brassica oleracea A. R 31.2 Triticosecale spp. R 33.7 Brassica oleracea A. R 31.4 Tropaeoluim maints R 1OO.O Brassica oleracea A. R 64.O Tropaeoluim maints O 31.5 Brassica oleracea A. S 68.7 Vaccinium anglisiifolium O 1OO.O Brassica oleracea A. R 75.3 Vaccinium anglisiifolium S 42.1 Brassica oleracea A. O 1OOO Vaccinium macrocarpon S 30.9 Brassica rapa A. S 27.6 Vicia viliosa R 35.5 Brassica rapa A. R 33.4 Vigna Sesquipedalis R 24.O Brassica rapa A. O 57.6 Vignatinguiculata R 31.6 Brassica rapa A. R S8.1 Vinca minor O 28.7 Brassica rapa A. R 84.5 Withania somnifera O 26.9 Calamintha nepeta A. O 6S.O Xanthium strumarium O 30.9 Cameia sinensis A. S 21.9 Zea mayS R 20.1 Cameia sinensis A. R 26.5 Zea mayS O 32.2 Cameia sinensis A. O 79.0 Cana eduis A. R 45.5 Canna eduis A. S 2O2 Capsella bursa-pastoris A. S 35.5 capsicum annuitin A. S 61.5 0245) capsicum annuitin A. O 89.8 capsicum annuitin A. R 1OOO TABLE 2 Capsicum finitesCens A. S 66.6 Capsicum finitesCens A. R 1OOO inhibition of MMP-2 by Plant Extracts Carthamus tinctorius A. R 21.3 Carthamus tinctorius A. R 21.5 Inhibition Chaerophyllum bulbosom A. R 57.2 Latin name Stress Extract (%) Chelidonium maius A. S 34.4 Chenopodium bonus-henricus A. R 43.5 Achillea millefolium A. S 21.9 Chenopodium bonus-henricus A. O 1OOO Achillea millefolium A. O 63.0 Chenopodium bonus-henricus A. R 76.4 Achillea millefolium A. R 1OOO Chenopodium quinoa A. O 92.0 Aconium napeius A. R 71.O Chrysanthemum coronarium A. R 48.6 Alcea rosea A. R 67.9 Chrysanthemum coronarium A. O 49.7 Aichemilia nois A. O 64.4 Chrysanthemun coronarium A. R 47.3 Allium ascalonictim A. R 20.9 Chrysanthemum coronarium A. R 26.7 Allium cepa A. R 84.3 Cicer arrieintin A. S 22.0 Allium grande A. R 36.7 Cicciarietinum A. O 23.6 Allium porrum A. O 1OOO Cichorium intybus A. S 21.1 Allium portin A. S 51.9 Cichorium intybus A. R 1OOO Allium portin A. R 66.7 Citritius ianatus A. S 65.5 Aium sativum A. R 1OOO Citritius ianatus A. R 96.3 Allium Schoenoprastin A. R 73.5 Citritius ianatus A. O 1OOO Aium Tiberosum A. S 24.3 Coix Lacryma-Jobi A. O 32.2 Aium Tiberosum A. O 83.6 Cornus Canadensis A. S 52.8 Aium Tiberosum A. R 89.3 Cosmos Sulphureus A. R 72.5 Aloe Vera A. R 69.7 Crataegus spp A. O 1OOO Althaea officinalis A. S 27.6 Cryptotaenia Canadensis A. R SO.6 Althaea officinalis A. R 64.7 Cryptotaenia Canadensis A. O 51.3 Amaranthus gangeticus A. S 29.4 Clictimis anguria A. S 53.4 Anethlin graveolens A. O 1OOO Clictimis Anguria A. R 84.9 Apium graveolens A. S 25.1 Cucumis meio A. R 91.7 Apium graveolens A. R 52.1 Cucurbita Maxina A. S 34.9 Aralia Cordata A. S 66.4 Cucurbita Maxina A. R 41.7 Aralia Cordata A. R 92.2 Cucurbita moschata A. R 36.8 Aralia nudicatiis A. O 29.4 Cucurbita moschata A. S 37.4 Arctii in minus A. S 28.4 Cucurbita pepo A. S 48.1 Armoracia rusticana A. S 2O2 Cucurbita pepo A. R 85.7 Armoracia rusticana A. O 55.0 Curcuna Zedoaria A. S 21.0 Arrhenaiherum eiatius A. S 40.2 Curcuna Zedoaria A. R 32.1 Artemisia dractinctius A. S 39.7 Curcurbita maxima A. S 27.0 Asparagus officinalis A. S 29.3 Cymbopogon citratus A. R 34.5 Atriplex hortensis A. R 33.6 Cymbopogon citratus A. O 1OOO Avena Saiiva A. R 37.2 Cymbopogon mariinii A. S 47.4 Beta vulgaris A. S 45.4 Dactylis glomerata A. S 20.6 Beta vulgaris A. R 95.9 Dactylis glomerata A. O 75.0 Beta vulgaris spp. Maritima A. R 1OOO Daiiciis Caroia A. S 445 Brassica chinensis A. R 49.6 Daiiciis Caroia A. R 70.5 US 2007/01 22492 A1 May 31, 2007 31

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Dipsacus sativus 40.4 Meilotus albus 1OOO Dirca palustris 27.2 Melissa officinalis 42.4 Doichos Labiah 54.2 Mentha pullegitim 88.3 Dryopteris filix-mas 76.3 Mentha spicaia 94.8 Echinacea purpurea 42.9 Mentha suaveoiens 82.9 Eleusine Coracana 37.5 Nepeta cataria 1OOO Eleusine Coracana 1OOO Nicotiana rustica 24.0 Erigeron Canadensis 45.7 Nicotiana rustica 1OOO Erica vesicaria 80.2 Nicotiana tabacum 42.5 Eschscholzia californica 42.4 Nicotiana tabacum 61.1 Eschscholzia californica 75.0 Nigella Saiiva 81.7 Eschscholzia californica 88.8 Ocimum tenuiflorum 23.1 Fagopyrim esculenium 1OOO Oenothera biennis 28.6 Fagopyri in tariarictim 38.6 Origantin majorana 52.9 Fagopyri in tariarictim 40.3 Origantin majorana 1OOO Fagopyri in tariarictim 71.O Origantim vulgare 66.8 Filipendula rubra 36.3 Panax quinquefoils 31.8 Foeniculum vulgare 41.6 Pastinaca Saiiva 27.7 Foeniculum vulgare 84.4 Pastinaca Saiiva 33.8 Foeniculum vulgare 1OOO Petasites japonicus 26.2 Forsythia intermedia 35.8 Petroselinum crispum 69.1 Fragaria X ananassa 97.2 Phaiaris canariensis 28.4 Gainsoga ciliata S4.O Phaiaris canariensis 29.7 Gaium odoratum 34.3 Phaiaris canariensis 94.3 Gaium odoratum 1OOO Phaseolus coccineus 30.8 Gaultheria hispidula 35.8 Phaseolus coccineus 79.5 Gaultheria hispidula 1OOO Phaseolus coccineus 80.9 Giaux maritima 46.5 Phaseolus mungo 59.8 Glycine max 27.0 Phaseolus vulgaris 47.3 Glycine Max 43.1 Phaseolus Vulgaris 744 Glycine max 1OOO Phaseolus vulgaris 83.2 Guizotia abyssinica 29.8 Phaseolus Vulgaris 1OOO Guizotia abyssinica 32.5 Phlox paniculata 23.7 Hamamelis virginiana 75.7 Phlox paniculata 81.7 Helianthus annuit is 69.0 Physalis alkekengi 23.5 Heianthus Tiiberosus 22.2 Physalis Ixocarpa 85.8 Heianthus tuberosus 69.7 Physalis ixocarpa 91.5 Heianthus Tiiberosus 1OOO Physalis Pruinosa 25.7 Hordeum hexastichon 22.3 Physalis Pruinosa 83.5 Hordeum hexastichon 34.9 Phytolacca decandra 31.5 Hordeum hexastichon 86.9 Phytolacca decandra 38.5 Hordeum vulgare 74.8 Pimpinella anisum 1OOO Hordeum vulgare subsp. Vulgare 34.5 Pimpinella anisum 1OOO Hordeum vulgare subsp. Vulgare 74.2 Plantago Coronopus 36.0 Hyssopus officinalis 57.5 Plantago Coronopus 38.4 Initia heienium 26.8 Plantago Coronopus S3.6 Ipomoea Batatas 20.1 Plantago major 65.3 Lathyrus Saivais 28.7 Plectranthus sp. 74.2 Lathyrus Saivais 1OOO Poa compressa 37.3 Lathyrus Sylvestris 42.4 Poa compressa 49.8 Lavandhiia iaiifolia 39.1 Poa compressa 1OOO Lepidium sativum 20.1 Polygontin pensylvanictim 63.5 Lepidium sativum 49.O Polygontin pensylvanictim 72.9 Levisticum officinale 23.0 Polygontin persicaria 27.5 Levisticum officinale 29.8 Polygontin persicaria 43.O Lintin itsitatissini in 56.9 Poterium sanguisorba 1OOO Lolium multiflorum 41.5 Poterium Sanquisorba 84.2 Lolium multiflorum 92.3 Pteridium aquilinum 45.1 Lotus corniculatus 95.5 Pteridium aquilinum 1OOO Lotus tetragonolobits 76.7 Pysalis ixocarpa 87.3 Lycopersicon esculenium 35.3 Raphantis raphanistrain 32.2 Lycopersicon esculenium 78.1 Raphantis Saivais 25.3 Lycopersicon esculenium 85.6 Raphantis Saivais 47.5 Lycopersicon pinpinoliifolium 74.9 Raphantis Saivais 83.5 Maiva moschata 21.5 Raphantis Saivais 84.7 Maiva moschata 445 Raphantis Saivais 1OOO Maiva verticiliata 22.0 Rheum officinale 44.0 Matricaria rectitita 40.9 Ribes nigrum 1OOO Matricaria rectitita 67.3 Ribes nigrum 1OOO Melaleuca alternifolia 6S.O Ricinits communis 1OOO Meilotus albus 50.7 Rosa rugosa 25.2 US 2007/01 22492 A1 May 31, 2007 32

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Ex 8. C t (%) Rosa rugosa 26.6 Vaccinium Corymbosum 1OOO Rosa rugosa 83.2 Veratrum viride 33.2 Rosmarinus officinalis 68.2 Verbascum thapsus 22.9 Rubus idaeus 819 Veronica beccabunga 52.8 Rubus idealis 734 Veronica officinalis 84.2 Rumex Acetosa 24.2 Vicia Saiva 1OOO Rumex Acetosa 85.5 Vicia viliosa 32.9 Rumex Acetosa 1OOO Vicia viliosa 1OOO Rumex crisputs 46.7 Vigna angularis S4.O Rumex crisputs 1OOO Vigna Sesquipedalis 48.3 Rita graveolens 1OOO Vigna Sesquipedalis 73.0 Saccharum officinarum 80.8 Vigna Sesquipedalis 96.6 Salix purpurea 56.7 Vignatinguiculata 70.7 Salvia officinalis 24.1 Vinca minor 22.1 Salvia officinalis 91.8 Vinca minor 88.4 Saivia Sciarea 99.7 Vitis sp. 20.9 Santolina Chamaecyparissus 83.8 Vitis sp. 3O4 Satureia hortensis 79.1 Xanthium Sibiricum 39.2 Satureia hortensis 1OOO Xanthium Sibiricum 47.8 Satureia montana 60.4 Xanthium Sibiricum 70.1 Satureia montana 76.1 Zea mayS 1OOO Scorzonera hispanica 22.1 Zea Mays 1OOO Secale cereale 47.2 Abelnochtis escientiis 21.6 Secale cereale 67.2 Abelnochtis escientiis 79.3 Senecio vulgaris 23.2 Achillea millefolium 62.7 Senecio vulgaris 76.6 Aconium napeius 82.O Sesamun indicum 1OOO Acorus calamus 1OOO Sesamun indicum 1OOO Ageratum conyzoides 49.3 Soianum duticamara 54.5 Alcea rosea 64.4 Soianum melanocerastin 45.4 Aichemia nois 21.5 Soianum melanocerastin 85.2 Aichemia nois 30.2 Soianum melanocerastin 88.7 Aichemia nois 55.7 Soianum melongena 42.5 Allium ampeioprastin 36.1 Soianum melongena 85.9 Allium ampeioprastin 52.8 Sonchi is oleracetis 25.6 Allium ascalonictim 68.9 Sorghum cafforum 39.6 Allium cepa 40.2 Sorghum dochna 3O.O Allium cepa 66.4 Sorghum dochna 48.0 Allium cepa 1OOO Sorghum dochna 62.O Allium grande 36.4 Sorghum durra 72.1 AAium sativum 29.5 Sorghum durra 94.6 AAium sativum 68.4 Sorghum Sudanense 1OOO AAium sativum 1OOO Spinacia oleracea 23.6 Allium Schoenoprastin 47.1 Stachys affinis 744 Allium Schoenoprastin 61.7 Stachys byzantina 48.4 Aium Tiberosum 23.8 Stachys byzantina 1OOO Aium Tiberosum 54.5 Stellaria graminea 20.8 Aium Tiberosum 85.9 Stellaria graminea 37.5 Aloe Vera S3.6 Steiaria media 49.O Althaea officinalis 37.4 Steiaria media 50.7 Altheaa officinalis 42.4 Symphytum officinale 44.2 Amaranthus caudathus 30.9 Tanacetum cinerariifolium 1OOO Amaranthus caudathus 56.7 Tanacetain parthenium 3O4 Amaranthus gangeticus 23.1 Tanacetum vulgare 28.6 Anethlin graveolens 23.9 Tanacetum vulgare 1OOO Angelica archangelica 22.0 Taraxacum officinale 59.1 Angelica archangelica 24.9 Thymus praecox. Subsp. arcticus 43.5 Apium graveolens 33.0 Thymus vulgaris 30.1 Apium graveolens 44.8 Thymus X citriiodorus 1OOO Apium graveolens 54.1 Trichosanthes kirilowii 29.2 Apium graveolens 84.1 Trichosanthes kirilowii 42.1 Aralia nudicatiis 51.8 Trigonelia foeningraeciin 53.4 Arctii in minus 25.4 Triticosecal spp. 44.8 Armoracia rusticana 52.1 Trictim aestivitin 65.5 Aronia melanocarpa 22.5 Triticum durum 53.9 Aronia melanocarpa 82.3 Triticum spelta 26.4 Artemisia dractinctius S3.6 Triticum spelta 36.7 Artemisia dractinctius 58.8 Triticum spelta 51.9 Artemisia dractinctius 1OOO Tropaeoluim maints 25.8 Artemisia dractinctius 1OOO Urtica dioica 22.9 Asclepias incarnata 26.9 Urtica dioica 30.6 Asparagus officinalis 24.0 US 2007/01 22492 A1 May 31, 2007 33

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Asparagus officinalis G R 65.9 Daiiciis Caroia G R 1OOO Asparagus officinalis G O 95.0 Digitalis purpurea G S 30.9 Aster spp G O 48.4 Dipsactis sativus G O 63.6 Beckmannia eruciformis G O 24.8 Dirca palustris G O 23.1 Bellis perennis G O 52.6 Doichos Labiah G S 33.0 Beta vulgaris G S 45.3 Dryopteris filix-mas G R 1OOO Beta vulgaris G R 1OOO Echinacea purpurea G R 93.4 Beta vulgaris spp. Maritima G R 1OOO Eleusine Coracana G S 3O.O Brassica cepicepa G R 52.9 Erigeron speciosus G S 28.9 Brassica chinensis G R 41.9 Errhenatherum elatius G S 55.6 Brassica iuncea G R 22.8 Erica vesicaria G R 54.7 Brassica naptis G S 22.9 Eschscholzia californica G S 47.9 Brassica oleracea G R 45.5 Eschscholzia californica G O 75.9 Brassica oleracea G R 47.1 Fagopyri in tartarictim G O 41.1 Brassica oleracea G S 62.9 Filipendula rubra G R 38.5 Brassica oleracea G R 77.9 Foeniculum vulgare G R 7O.O Brassica oleracea G O 1OOO Foeniculum Vulgare G S 1OOO Brassica rapa G S 26.5 Gainsoga ciliata G S 34.6 Brassica rapa G R 38.9 Gainsoga ciliata G R 48.2 Brassica rapa G R S3.6 Gaultheria hispidula G R 6O.S Calamintha nepeta G S 20.4 Gaultheria hispidula G O 1OOO Calamintha nepeta G O 78.0 Gaultheria hispidula G S 1OOO Cameia sinensis G O 1OOO Giaux maritima G R 59.3 Campaniiia raptinctiitis G R 60.6 Glycine max G R 21.1 Canna editiis G O 78.1 Glycine max G S 24.4 Capsella bursa-pastoris G S 30.7 Glycine max G O 28.1 Capsella bursa-pastoris G R 60.6 Guizotia abyssinica G S 26.0 capsicum annuum G S 70.8 Guizotia abyssinica G R 36.8 capsicum annuitin G O 8O.O Guizotia abyssinica G O 1OOO capsicum annuitin G R 1OOO Hedeona pullegioides G O 94.6 Capsicum finitesCens G S 63.2 Helianthus anniitis G S 35.5 Capsicum finitesCens G R 1OOO Helianthus anniitis G O 75.0 Carthamus tinctorius G R 1OOO Helianthus anniitis G R 79.9 Centatirea solstitialis G S 46.4 Heianthus strumosus G O 1OOO Cerasitim to mentostin G R 52.3 Heianthus tuberosus G R 642 Chenopodium bonus-henricus G S 22.0 Helichrysum thianschanicum G O 61.1 Chenopodium quinoa G S 31.0 Helleborus niger G R 48.0 Chenopodium quinoa G O 53.4 Hordeum hexastichon G S 26.8 Chrysanthemun coronarium G R 76.2 Hordeum vulgare G O 65.4 Chrysanthemum coronarium G R 54.2 Hordeum vulgare subsp. Vulgare G O 75.8 Cicer arrieintin G S 23.1 Huntilus lupulus G S 26.0 Cichorium endivia subspendivia G S 28.7 Hypericum hennyi G R 2O2 Cichorium endivia subspendivia G O 68.7 Hypericum hennyi G O 71.1 Cichorium intybus G S 41.4 Hyssopus officinalis G O 1OOO Cichorium intybus G O 62.1 Iberis amara G S 21.2 Circium arvense G S 25.3 Initia heienium G S 24.3 Circium arvense G R 59.3 Lactica Saiiva G R 1OOO Citritius ianatus G S 24.8 Lactuca serioia G R 69.3 Citritius ianatus G R 41.1 Laportea Canadensis G R 1OOO Citritius ianatus G R 1OOO Lathyrus Sylvestris G O 39.6 Cosmos Sulphureus G R 77.9 Lavandhiia angustifolia G O 7O.O Cosmos Sulphureus G S 79.4 Lavandhiia iaiifolia G S 22.7 Ciclinis Saiivus G S 39.9 Lepidium Sativum G R 30.6 Ciclinis Saiivus G S 39.9 Lepidium sativum G S 53.3 Cucurbita maxima G S 33.9 Levisticum officinale G O 80.7 Cucurbita maxima G R 43.4 Lolium multiflorum G O 34.5 Cucurbita maxima G O 1OOO Lotus corniculatus G S 32.9 Cucurbita moschata G S 41.3 Lotus corniculatus G O 1OOO Cucurbita pepo G S 42.8 Lotus tetragonolobits G R 79.9 Cucurbita pepo G S 45.4 Lycopersicon escientiin G S 28.2 Cucurbita Pepo G R 83.0 Lycopersicon escientiin G R 75.4 Cuminum cyminum G O 66.2 Lycopersicon pinpinellifolium G R 81.4 Curcuna Zedoaria G R 33.9 Maius hupehensis G R 32.5 Cymbopogon citratus G R 65.8 Maius hupehensis G S 41.2 Cymbopogon mariinii motia G S 41.4 Maiva moschata G O 47.1 Cymbopogon mariinii motia G O 6O.S Malva Sylvestris G S 23.1 Dactylis glomerata G S 21.9 Maiva verticiliata G R 39.9 Dactylis glomerata G O 61.2 Matricaria recuita G O 3O.O Daitira Stranonium G S 27.0 Matricaria recuita G S 71.3 Daiiciis Caroia G O 21.3 Melaleuca alternifolia G O 58.3 Daiiciis Caroia G S 31.0 Meilotus aiba G S 41.1 US 2007/01 22492 A1 May 31, 2007 34

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Meilotus albus 88.8 Poa pratensis 28.2 Meilotus albus 1OOO Polygonum aviculare 1OOO Melissa officinalis 47.8 Polygontin pensylvanictim 27.7 Mentha arvensis 33.9 Polygontin pensylvanictim 54.1 Mentha arvensis 6.3.3 Polygontin persicaria 32.O Mentha piperita 32.3 Polygontin persicaria 35.7 Mentha piperita 85.9 Polygontin persicaria 1OOO Mentha piperita 1OOO Portuiaca oleracera 51.5 Mentha spicaia 28.9 Poterium sanguisorba 89.9 Mentha spicaia 37.5 Poterium sanguisorba 1OOO Mentha suaveolens 25.6 Poterium sanquisorba 23.7 Mentha suaveolens 70.3 Prunella vulgaris 26.7 Monoraica charantia 52.9 Prunus cerasifera 95.3 Monarda didyma 22.0 Raphantis Raphanistrain 41.7 Monarda didyma 1OOO Raphantis Raphanistrain 43.5 Monarda fistulosa 26.0 Raphantis Saivais 41.O Nepeta cataria 23.4 Raphantis Saivais 44.6 Nicotiana tabacum 45.2 Raphantis Saivais 50.5 Nigella Saiiva 94.7 Raphantis Saivais 86.1 Ocimum basicum 23.0 Raphantis Saivais 1OOO Ocimum basicum 1OOO Reseda Odorata 58.3 Ocimum tenuiflorum 45.3 Rheum officinale 30.7 Oerothera biennis S4.3 Ribes nigrum S4.3 Origantin majorana 1OOO Ribes nigrum 63.8 Origantin majorana 1OOO Ribes Sylvestre 1OOO Origantim vulgare 93.3 Ricinits communis 41.5 Origantim vulgare 93.5 Ricinits communis 1OOO Origanum vulgare 97.4 Rosmarinus officinalis 90.0 Oxalis Deppei 28.7 Rubus idaeus 37.1 Oxalis Deppei 87.2 Rubus idealis 26.6 Oxalis Deppei 1OOO Rubus occidentais 35.1 Oxyria digyna 54.5 Rumex crispiis 30.3 Panicum miliaceum 71.1 Rumex crispiis 1OOO Panicum miliaceum 1OOO Rumex patientia 41.O Panicum miliaceum 1OOO Rumex patientia 41.9 Passiflora caerula 26.3 Rita graveolens 47.9 Passiflora caerula 72.1 Rita graveolens 82.1 Pastinaca Saiiva 24.3 Saccharum officinarum 1OOO Pastinaca Saiiva 90.2 Salvia elegens 1OOO Petroselinum crispum 87.6 Salvia officinalis 35.3 Petroselinum crispum 1OOO Salvia officinalis 1OOO Phaiaris canariensis 1OOO Salvia officinalis 1OOO Phaiaris canariensis 1OOO Sambucus ebulus 53.9 Phaseolus acutifolius 79.6 Santolina Chamaecyparissus 36.4 Phaseolus coccineus 28.3 Santolina Chamaecyparissus 69.5 Phaseolus coccineus 804 Santolina Chamaecyparissus 1OOO Phaseolus mingo 37.2 Saponaria officinalis 29.8 Phaseolus vulgaris S4.3 Satureia hortensis 97.4 Phaseolus vulgaris 59.0 Satureia hortensis 1OOO Phaseolus vulgaris 73.7 Satureia montana 59.2 Phaseolus vulgaris 1OOO Satureia repandra 35.3 Phlox paniculata 37.7 Satureia repandra 66.2 Phlox paniculata 77.0 Scorzonera hispanica 24.5 Phlox paniculata 80.8 Scrophtiaria nodosa 24.5 Physalis ixocarpa 3O.S Scrophtiaria nodosa 3O.O Physalis ixocarpa 78.3 Scrophtiaria nodosa 55.6 Physalis ixocarpa 80.9 Scutellaria laterifiora 20.3 Physalis pruinosa 63.2 Scutellaria laterifiora 83.1 Phytolacca americana 36.1 Secale cereale 51.1 Phytolacca americana 1OOO Senecio vulgaris 42.5 Pimpinella anisum 26.1 Sesamun indicum 34.3 Pimpinella anisum 3O.O Sesamun indicum 445 Pistin Saivitin 28.4 Silene vulgaris 34.1 Plantago Coronopus 27.8 Siim Sisartin 1OOO Plantago Coronopus 51.1 Soianum melanocerastin 40.6 Plantago Coronopus 67.5 Soianum melanocerastin 85.4 Plantago major 30.3 Soianum melongena 58.2 Plantago major 64.6 Soianum melongena 83.0 Poa compressa 63.0 Soianum melongena 85.6 Poa compressa 674 Soianum tuberosum 40.2 Poa compressa 89.0 Sonchi is oleracetis 41.1 US 2007/01 22492 A1 May 31, 2007 35

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Ex 8. C t (%) Latin name Extract (%) Sorghum dochna 2SO Perilla frutescens 1OOO Sorghum dochna 64.3 Abies lasiocarpa 2O2 Sorghum dochna 1OOO Abies lasiocarpa 59.1 Sorghum dirra 60.1 Achillea millefolium 84.7 Sorghum durra 1OOO Aconium napeius 22.0 Sorghum Sudanense 98.0 Aconium napeius 1OOO Spinacia oleracea 24.9 Adiantum pedatum 1OOO Spinacia oleracea 1OOO Agarict is bisports 52.1 Stachys byzantina 78.8 Agarict is bisports 65.6 Stellaria graminea 29.3 Ageratum conyzoides 26.7 Steiaria media 33.4 Agropyron repens 30.2 Steiaria media 45.4 Agrostis Stoionifera 1OOO Symphytum officinale 57.5 Alcea rosea 63.7 Tanacetum cinerariifolium 1OOO Aichemia nois 28.6 Tanacetain parthenium 28.2 Allium ampeioprastin 55.9 Tanacetum vulgare 25.2 Allium ampeioprastin 60.4 Tanacetum vulgare 39.3 Allium ascalonictim 20.4 Tanacetum vulgare 81.2 Allium ascalonictim 734 Taraxacum officinale 51.1 Allium cepa 33.8 Thymus fragantissimils 29.9 Allium cepa 35.6 Thymus fragantissimils 55.3 Allium cepa 48.0 Thymus praecox. Subsp. arcticus 27.7 Allium cepa 78.6 Thymus serpyllum 74.9 Allium grande 32.4 Thymus vulgaris 23.3 Allium Schoenoprastin 67.7 Thymus vulgaris 86.4 Aium Tiberosum 38.8 Thymus X citriiodorus 97.6 Aium Tiberosum 82.5 Tragopogon porrifolius 76.2 Aium Tiberosum 85.2 Trichosanthes kirilowii 87.7 Aloe Vera 74.6 Trigonelia foeningraeciin 31.0 Althaea officiana is 37.7 Trigonelia foeningraeciin 84.O Althaea officinalis 55.3 Triticosecale spp 26.5 Althaea officinalis 72.3 Triticosecale spp 73.5 Amaranthus caudathus 53.5 Trictim aestivitin 624 Amaranthus gangeticus 28.1 Triticum durum 51.9 Ananas Conostis 37.9 Triticum spelta 24.5 Ananas Conostis 1OOO Triticum spelta 32.9 Angelica archangelica 41.3 Triticum turgidium 25.1 Anthemis nobiis 1OOO Tropaeoluim maints 21.3 Anthemis nobiis 1OOO Tropaeoluim maints 45.6 Anthriscus cerefolium 21.9 Urtica dioica 21.3 Anthriscus cerefolium 67.1 Urtica dioica 1OOO Apium graveolens 35.5 Valerianeia iocusia 32.2 Apium graveolens 52.1 Veratrum viride 777 Aralia Cordata 1OOO Verbascum thapsus 34.0 Aralia nudicatiis 31.2 Veronica beccabunga 44.1 Arctii in minus 31.3 Veronica officinalis 38.8 Arctii in minus 73.7 Veronica officinalis 87.5 Armoracia rusticana 49.9 Viburnum trilobium 62.6 Arrhenatherum elatius 1OOO Vicia faba 22.2 Artemisia dractinius 1OOO Vicia Saiva 74.8 Asclepias incarnata 32.3 Vicia Saiva 1OOO Asparagus officinalis 48.2 Vicia viliosa 1OOO Atriplex hortensis 28.4 Vigna angularis 65.2 Avena Saiiva 31.3 Vigna Sesquipedalis 35.1 Avena Saiiva 70.6 Vigna Sesquipedalis 73.8 Avena Saiiva 1OOO Vigna Sesquipedalis 1OOO Averrhoa caramboia 44.0 Vignatinguiculata 65.9 Bellis perennis 82.O Vignatinguiculata 84.5 Beta vulgaris 33.7 Vinca minor 22.1 Beta vulgaris 1OOO Vitis sp. 40.1 Betula glandulosa 53.5 Vitis sp. 74.7 Boietus eduis 21.8 Withania somnifera 37.3 Borago officinalis 423 Withania somnifera 91.0 Borago officinalis 78.5 Xanthium Sibiricum 38.4 Brassica hiria 53.1 Xanthium Sibiricum 1OOO Brassica hiria 68.9 Xanthium strumarium 37.7 Brassica naptis 45.1 Xanthium strumarium 39.6 Brassica naptis 82.9 Xanthium strumarium 40.O Brassica oleracea 38.8 Zea mayS 43.3 Brassica oleracea 49.7 Zea mayS 64.4 Brassica oleracea 75.5 Zea mayS 68.3 Brassica oleracea 77.0 US 2007/01 22492 A1 May 31, 2007 36

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Ex 8. C t (%) Latin name Extract (%) Brassica oleracea 77.2 Cucurbita pepo 45.8 Brassica rapa 25.4 Cucurbita pepo 80.2 Brassica rapa 37.9 Cucurbita pepo 98.9 Brassica rapa 47.7 Cuminum cyminum S4.O Brassica rapa 64.7 Curcuna Zedoaria 1OOO Brassica rapa 81.8 Cymbopogon citratus 21.0 Calamintha nepeta 57.6 Cymbopogon mariinii motia 27.5 Calendula officinalis 32.6 Cynara Scolymits 23.1 Cameia sinensis 21.0 Cynara Scolymits 834 Cameia sinensis 43.8 Cypertis esculenius 1OOO Cameia sinensis 66.2 Dactiis Giomerata 30.8 Canna editiis 1OOO Dactiis Giomerata 34.5 Canihareilus cibarias 26.0 Daiiciis Caroia 27.1 capsicum annuitin 54.6 Daiiciis Caroia 56.8 capsicum annuitin 1OOO Daiiciis Caroia 1OOO Capsicum finitesCens 60.9 Digitalis purpurea 38.4 Capsicum finitesCens 1OOO Dirca palustris 45.9 Carex morrowii 24.4 Doichos iabiab 46.6 Carica papaya 20.8 Dryopteris filix-mas 29.5 Carthamus tinctorius 39.6 Dryopteris filix-mas 1OOO Carya cordiformis 1OOO Echinacea purpurea 59.3 Cerasitim to mentostin 54.8 Echinacea purpurea 87.8 Chaerophyllum bulbosum 42.2 Eleusine Coracana 28.6 Chaerophyllum bulbosum 74.3 Eleusine Coracana 8O.O Chelidonium maius 20.3 Erigeron Canadensis 1OOO Chenopodium quinoa 76.O Erica vesicaria 6O.S Chrysanthemum coronarium 30.6 Erysimum perofskianum 28.2 Chrysanthemum parthenium 57.2 Erysimum perofskianum 85.2 chrysanthemun coronarium 56.5 Eschscholzia californica 49.9 Chrysanthemum coronarium 81.6 Eschscholzia californica 74.5 Cicer arrieintin 32.2 Fagopyrim esculenium 52.9 Cichorium endivia subspendivia 27.1 Fagopyri in tartarictim 25.6 Cichorium endivia subsp. Endivia 26.9 Fagopyri in tartarictim 68.4 Cichorium endivia subsp. Endivia 64.5 Fagopyri in tartarictim 1OOO Cichorium intybus 22.7 Festica rubra 51.6 Cichorium intybus 53.5 Festica rubra 56.6 Cinicifiiga racemosa 41.1 Festica rubra 71.7 Cinicifiiga racemosa 68.4 Foeniculum vulgare 36.5 Circium arvense 42.5 Foeniculum vulgare 41.4 Circium arvense 64.5 Foericulum vulgare 1OOO Citritius ianatus 724 Fortunella spp 53.9 Citritius ianatus 92.2 Fragaria X ananassa 28.1 Citritius ianatus 1OOO Gainsoga ciliata 43.2 Citrus innettoides 77.1 Gainsoga ciliata 73.3 Citrus innon 43.6 Gaium odoratum 42.O Citrus paradisi 21.8 Gaium odoratum 94.2 Citrus paradisi 90.9 Giatix Maritima 24.8 Citrus Sinensis 46.7 Glycine max 37.2 Colocasia sp 43.4 Glycine max 1OOO Colocasia sp 84.3 Glycine max 1OOO Corchorus oitorius 22.7 Glycine max 1OOO Coriandrum sativum 20.4 Gossypium herbaceum 48.7 Cornus Canadensis 66.O Guizotia abyssinica 26.8 Cosmos Sulphureus 47.1 Guizotia abyssinica 1OOO Crataegus submolis 21.2 Hedeona pullegioides 20.3 Crataegus submolis 94.3 Hedeona pullegioides 72.7 Cuctimis anguria 49.4 Helianthus anniitis 56.1 Cuctimis anguria 84.1 Heianthus strumosus 1OOO Cucumis meio 56.6 Heianthus tuberosus 25.3 Cucumis meio 92.4 Heianthus tuberosus 28.1 Cucumis meio 1OOO Heianthus tuberosus 78.6 Cucumis metuliferus 29.5 Heianthus tuberosus 91.5 Ciclinis Saiivus 28.3 Helichrysum angustifolium 834 Cucurbita maxima 26.7 Helichrysum angustifolium 88.3 Cucurbita maxima 34.7 Helichrysum thianschanicum 26.0 Cucurbita maxima 62.1 Heliotropium arborescens 1OOO Cucurbita moschata 30.7 Helleborus niger 23.0 Cucurbita moschata 33.4 Hibiscus cannabinus 37.9 Cucurbita moschata 48.3 Hordeum vulgare 75.9 Cucurbita moschata 98.8 Hordeum vulgare supsp vulgare 2O.S Cucurbita moschata 1OOO Hordeum vulgare supsp vulgare 62.3 US 2007/01 22492 A1 May 31, 2007 37

TABLE 2-continued TABLE 2-continued inhibition of MMP-2 by Plant Extracts inhibition of MMP-2 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Huntilus lupuits S 44.7 Oenothera biennis R 48.2 Huntilus lupuits O 70.6 Onobrychis viciifolia S 34.4 Hypericum henryi O 76.8 Onobrychis viciifolia O 35.6 Hypericum henryi R 99.8 Opiiniia sp. S 23.5 Hyperictim perforatin R 38.8 Origantim vulgare S 20.7 Hyssopus officinalis O 1OOO Origantim vulgare R 76.7 Iberis amara O 1OOO Origantim vulgare O 1OOO Juniperus Communis S 1OOO Oryza sativa R 60.8 Kochia scoparia S 25.2 Oxalis Deppei S 22.2 Koelleria giatica S 23.1 Oxalis Deppei R 81.4 Lactica Saiiva R 70.5 Passiflora caerulea S 36.9 Lactuca serioia R 34.1 Passiflora caerulea R 87.O Laportea Canadensis R 61.3 Passiflora spp R 54.6 Lathyrus Sylvestris R 48.6 Pastinaca Saiiva S 24.8 Latiri is nobilis O 73.6 Pastinaca Saiiva R 74.7 Lavandhiia angustifolia R 3S.O Perroselinum crispum R 85.2 Lavandhiia angustifolia O 1OOO Perroselinum crispum O 1OOO Lavandhiia iaiifolia O 77.1 Persea americana R 43.1 Lepidium sativum S 35.2 Petasites Japonicus S 21.9 Lepidium sativum R 48.1 Petroselinum crispum R 52.8 Lepidium sativum O 72.9 Peucedanum or'easelinum R 41.9 Levisticum officinale S 38.7 Phaiaris canariensis R 41.1 Levisticum officinale O 60.3 Phaiaris canariensis O 1OOO Lintin itsitatissini in R 24.7 Phaseolus acutifolius R 88.2 Lolium multiflorum S 39.8 Phaseolus coccineus S 22.2 Lolium multiflorum O 74.1 Phaseolus coccineus R 36.4 Lonicera ranosissina S 34.4 Phaseolus coccineus R 86.7 Lonicera ranosissina O 8O.S Phaseolus coccineus O 1OOO Lonicera syringantha R 58.4 Phaseolus mingo S 43.O Lotus corniculatus S 36.0 Phaseolus vulgaris S 62.9 Lotus corniculatus O 1OOO Phaseolus vulgaris R 71.9 Lotus tetragonolobits R 76.1 Phaseolus vulgaris R 73.0 Linaria annia R 47.4 Phaseolus vulgaris O 1OOO Lycopersicon esculenium R 69.7 Phlox paniculata R 23.1 Lycopersicon pinpineliifolium R 58.7 Phlox paniculata R 92.8 Maius hupehensis R 53.1 Physalis alkekengi R 39.5 Maius hupehensis S 1OOO Physalis ixocarpa R 36.7 Maius sp. R 72.6 Physalis ixocarpa R 75.9 Maiva moschata O 96.7 Physalis pruinosa R 65.6 Maiva verticiliata R 35.8 Physalis pruinosa R 71.O Manihot escuienia R 53.7 Physalis pruinosa O 1OOO Melaleuca alternifolia S 21.5 Physalis pruinosa O 1OOO Melaleuca alternifolia O 78.7 Phytolacca decandra S 39.3 Meilotus albus R 79.7 Phytolacca decandra O 42.O Melilotus officinalis S 34.6 Pimpinella anisum S 27.9 Melilotus officinalis R 1OOO Pimpinella anisum R 35.8 Melissa officinalis O 1OOO Pimpinella anisum O 49.9 Mentha piperita S 24.5 Pimpinella anisum R 55.5 Mentha pullegitim O 1OOO Pistin Saivitin S 22.3 Mentha suaveolens O 20.9 Plantago Coronopus R 35.2 Miscanthus sinensis Andress S 69.1 Plantago Coronopus R 46.0 Monoraica charantia R S4.9 Plantago Coronopus O 73.5 Monarda didyma S 31.3 Plantago major S 22.3 Monarda fistulosa S 21.3 Plectranthus sp. S 59.2 Monarda fistulosa O 1OOO Pleurotus spp R 26.6 Montia perfoliata R 67.2 Poa compressa S 33.4 Musa paradisiaca R 47.3 Poa compressa R 75.7 nastairtium officinale S 55.7 Poa compressa O 1OOO Nepeta cataria S 20.7 Poa pratensis S 25.4 Nepeta cataria S 69.0 Polygontin pensylvanictim O 66.8 Nepeta cataria O 1OOO Polygontin pensylvanictim R 73.3 Nicotiana rustica S 52.8 Polygontin persicaria S 27.1 Nicotiana rustica R 88.1 Polygontin persicaria O SO.8 Nicotiana tabacum S S.O.3 Populus incrassata O 74.3 Nicotiana tabacum R 91.5 Populus incrassata S 1OOO Nigella Saiiva R 34.2 Prints armeniaca R 55.0 Nigella Saiiva R 90.3 Prints cerastis O 1OOO Nigella Saiiva R 1OOO Prunus persica S 26.0 Ocimum Basilicum S 21.6 Prunus persica R 46.2 Ocimum Basilicum O 1OOO Psoralea corylifolia S 47.4 Ocimum tenuiflorum R 445 Pteridium aquilinum R 1OOO

US 2007/01 22492 A1 May 31, 2007 39

TABLE 2-continued TABLE 3-continued inhibition of MMP-2 by Plant Extracts Inhibition of MMP-3 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%)

Xanthium strumarium T O 74.3 Fragaria X (SS A. S 67.1 Zea mayS T R 55.7 Fragaria X (SS A. O 99.6

Zea mayS T O 1OOO eCEACEattineria nispianita 5. Zingiber officinale T R 79.0 Glycyrrhiza glabra A. R 56.2 Hedeona pullegioides A. O 51.7 Heianthus tuberosus A. O 22.9 0246 Hordeum vulgare subsp. vulgare A. S 36.0 Hypericum hennyi A. R 67.2 Hyperictim perforatin A. R 31.7 TABLE 3 Hyssopus officinalis A. R 21.6 Iris versicolor A. R S3.6 Inhibition of MMP-3 by Plant Extracts Isais inctoria A. S 32.9 Levisticum officinale A. O 46.7 Inhibition Lotus tetragonolobits A. R 26.2 Latin name Stress Extract (%) Matricaria recuita A. S 43.5 Matteucia pensylvanica A. R 24.7 Achillea millefolium A. O 21.4 Melissa officinalis A. S 30.3 Aium Tiberosum A. S 32.5 Mentha suaveoiens A. R 91.7 Anethlin graveolens A. S 26.0 Nepeta cataria A. S 30.3 Anthemis nobiis A. R 20.3 Nigella Saiiva A. O 26.0 Anthemistinctoria A. R 58.0 Ocinum tenuiflorum A. O 33.0 Apium graveolens A. R 34.1 Ocinum tenuiflorum A. R 49.8 Arctii in minus A. R 53.9 Perilla frutescens A. R 34.8 Arctii in minus A. O 1OOO Petasites japonicus A. R 38.0 Arctostaphylos tiva-ursi A. S 58.6 Phaseolus mingo A. O 62.6 Aronia melanocarpa A. R 32.2 Phaseolus vulgaris A. S 21.2 Artemisia Absinthium A. O 1OOO Phaseolus vulgaris A. O SO.6 Artemisia dractinctius A. R 23.4 Phaseolus Vulgaris A. R 1OOO Artemisia dractinctius A. S 63.0 Phlox paniculata A. S 46.4 Aster sp A. O 42.4 Physalis alkekengi A. O 37.5 Atropa belladonna A. O 23.8 Plantago major A. O 27.3 Beta vulgaris A. S 24.1 Polygonum aviculare line A. S 24.8 Beta vulgaris A. O 42.9 Polygontin persicaria A. S 59.1 Beta vulgaris A. O 94.3 Potentiiia anserina A. R 40.1 Beta vulgaris A. R 97.9 Poterium sanguisorba A. R 75.7 Beta vulgaris var. Condiwata A. O 21.2 Prunus cerasifera A. R 8O.O Brassica naptis A. S 2SO Piaridium aquilinus A. R 39.6 Brassica naptis A. O 1OOO Raphantis raphanistrain A. S 28.2 Brassica oleracea A. S 39.9 Raphantis Saivais A. S 64.4 Canna editiis A. S 39.6 Ribes nigrum A. O 47.6 Capsicum annittin A. S 35.4 ribes tiva-Crispa A. R 21.0 Capsicum finitesCens A. S 27.2 ribes tiva-Crispa A. O 1OOO Cichorium intybus A. O 2O2 Rosa rugosa A. S 21.4 Cichorium intybus A. R 26.5 Rosmarinus officinalis A. R 27.3 Cichorium intybus A. S 28.2 Rubus allegheniensis A. R 81.0 Citritius ianatus A. S 21.7 Rubus arcticus A. R 51.0 Citritius ianatus A. O 27.8 Rubus Canadensis A. R 48.8 Citritius ianatus A. R 34.4 Rubus idaeus A. S 28.5 Coix Lacryma-Jobi A. S 37.3 Rubus idaeus A. R 35.1 Coix Lacryma-Jobi A. O 78.1 Rubits pubescens A. O SO4 Cosmos Sulphureus A. R 26.8 Rubus thibetanus A. O 39.1 Crataegus submolis A. S 22.3 Rumex patientia A. S 24.8 Crataegus submolis A. R 61.6 Rita graveolens A. O 56.1 Cuctimis anguria A. S 27.8 Salvia officinalis A. R 43.2 Cucurbita Maxina A. S 28.9 Santolina Chamaecyparissus A. R 27.0 Cucurbita moschata A. S 32.9 Scutellaria laterifiora A. R 53.5 Cucurbita pepo A. S SO.9 Soianum melongena A. S 21.8 Datisca Cannabina A. R 43.3 Solidago Canadensis A. S 27.4 Datisca Cannabina A. S 1OOO Stachys affinis A. S 1OOO Digitalis purpurea A. R 2O.O Steiaria media A. O 24.4 Dipsacus sativus A. R 64.8 Tanacetum vulgare A. R 62.1 Dirca palustris A. S 29.6 Thymus praecox. Subsp. arcticus A. S 28.4 Dryopteris filix-mas A. R 22.0 Thymus praecox. Subsp. arcticus A. O 31.8 Dryopteris filix-mas A. O 32.8 Trichosanthes kirilowii A. S 23.2 Echinacea purpurea A. O 1OOO Vaccinium Corymbosum A. R 1OOO Fagopyri in tatarictim A. R 28.3 Vaccinium macrocarpon A. S 48.6 Fagopyri in tatarictim A. O 29.7 Vaccini in augustifolium A. R 56.6 Filipendula rubra A. S 43.7 Vigna angularia A. O 23.1 Filipendula rubra A. R 63.2 Vigna Sesquipedalis A. O 37.8 Fragaria X ananassa A. R 41.5 Vignatinguiculata A. S 52.5 US 2007/01 22492 A1 May 31, 2007 40

TABLE 3-continued TABLE 3-continued Inhibition of MMP-3 by Plant Extracts Inhibition of MMP-3 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Vinca minor A. O 23.2 Malva Sylvestris G O 73.7 Vitis sp. A. S 20.8 Matricaria recuita G S 31.5 Vitis sp. A. O 21.5 Melaleuca alternifolia G S 21.5 Vitis sp. A. R 33.6 Melissa officinalis G S 32.8 Xanthium Sibiricum A. S 27.3 Melissa officinalis G R 44.8 Aconium napeius G O 59.0 Melissa officinalis G O 82.4 Agropyron repens G O 694 Mentha piperita G R 77.3 Aichemilia nois G S 30.6 Mentha pullegitim G R 41.1 Aichemilia nois G O 73.3 Monarda didyma G S 31.8 Allium grande G O 33.4 Nepeta cataria G R 25.8 Anethlin graveolens G S 4.O.S Nepeta cataria G O 84.9 Aronia melanocarpa G O 1OOO Nigella Saiiva G O 44.9 Artemisia absinthium G S 31.3 Ocinum tenuiflorum G R 23.7 Artemisia absinthium G O 67.9 Oenothera biennis G S 25.6 Artemisia dractinctius G S 1OOO Origantim vulgare G S 28.6 Atropa belladonna G S 41.2 Origantim vulgare G R 31.2 Bellis perennis G S 48.4 Pennisetum alopectiroides G S 49.9 Brassica oleracea G S 26.4 Petroselinum crispum G S 31.5 Brassica oleracea G O 40.6 Peucedanum or'easelinum G R 68.3 Brassica rapa G S 21.4 Phaseolus acutifolius G R 25.4 Capsicum annittin G S 3S.O Phaseolus acutifolius G O 618 Capsicum annittin G S 35.7 Phaseolus vulgaris G O 24.4 Capsicum finitesCens G S 27.5 Phaseolus vulgaris G S 35.6 Chelidonium maius G O 34.7 Phlox paniculata G S 27.2 Cichorium intybus G R 34.4 Physalis alkekengi G R 26.1 Coix Lacryma-Jobi G S 2O2 Physalis alkekengi G O S4.9 Cosmos Sulphureus G O 32.9 Plantago major G O 55.9 Crataegus submolis G S 25.6 Plectranthus sp. G R 23.0 Crataegus submolis G R 28.6 Polygontin persicaria G S 41.1 Cuctimis anguria G S 33.6 Potentiiia anserina G R SS.4 Cucurbita maxima G S 44.6 Poterium sanguisorba G R 76.4 Cucurbita moschata G S 33.4 Prunus cerasifera G R 55.3 Cucurbita pepo G S 25.3 Piaridium aquilinus G R 445 Cymbopogon citratus G S 30.3 Rhaphant is sativus G O 98.1 Cymbopogon mariinii G S 61.1 Rheum x cuitorum G R 27.0 Daiiciis Caroia G O 3O.O Ribes nidigrolaria G R 22.0 Dryopteris filix-mas G S 26.0 Ribes Silvestris G R 88.8 Dryopteris filix-mas G R 45.3 Rosmarinus officinalis G R 39.4 Echinacea purpurea G O 51.8 Rubus idaeus G S 1OOO Echinochloa frumentacea G S 30.3 Rubus idealis G O 37.0 Fagopyrim esculenium G R SO.9 Rubus Phoenicaiasius G R 24.9 Fagopyri in tariarictim G O 44.0 Rubits pubescens G O 23.0 Fagopyri in tariarictim G R 46.0 Rubus thibetanus G O 41.2 Filipendula rubra G S 53.1 Rumex patientia G S 36.2 Filipendula rubra G R 58.7 Salvia officinalis G O 34.5 Forsythia intermedia G O 52.9 Salvia officinalis G R 89.5 Fragaria X ananassa G R 40.7 Sanguisorba officinalis G S 46.8 Fragaria X ananassa G R 28.1 Santolina Chamaecyparissus G R 33.7 Gaultheria hispidula G R 72.8 Secale cereale G S 24.4 Gaultheria hispidula G O 1OOO Senecio vulgaris G R 37.6 Gaultheria procumbens G R 24.1 Soianum melongena G S 21.1 Glycine max G S 31.2 Soianum tuberosum G S 27.6 Glycyrrhiza glabra G R 37.1 Sorghum dochna G S 23.7 Guizotia abyssinica G R 35.4 Sorghum dochna G R 56.3 Hamamelis virginiana G S 29.1 Symphytum officinale G S 25.2 Hamamelis virginiana G R 67.1 Teucrium chamaedrys G S 75.4 Helenium hoopesii G R 39.8 Thymus praecox. Subsp. arcticus G S 28.4 Heianthus tuberosus G O 32.8 Thymus praecox. Subsp. arcticus G O 52.1 Hordeum hexastichon G S 60.9 Thymus X citriiodorus G R 25.3 Huntilus lupuits G R 61.2 Triticum durum G S 21.9 Huntilus lupuits G S 90.5 Triticum turgidium G O 80.2 Hypericum henryi G R 1OOO Vaccinium anglisiifolium G R 47.6 Hyperictim perforatin G R 43.4 Vaccinium anglisiifolium G R 48.1 Hyssopus officinalis G S 25.1 Vaccinium anglisiifolium G R 71.O Hyssopus officinalis G O 48.2 Vaccinium corymbosum G R 60.6 Iris versicolor G R 47.0 Vaccinium corymbosum G R 61.7 Isais inctoria G S 32.1 Vaccinium corymbosum G O 99.4 Lavandhiia angustifolia G S 43.9 Vaccinium macrocarpon G R 1OOO Levisticum officinale G O 514 Vaccini in anguistifolium G O 24.4 Maius hupehensis G S 24.2 Vaccini in anguistifolium G R 41.5 Maius hupehensis G R 37.2 Valeriana oficinalis G R 33.5 US 2007/01 22492 A1 May 31, 2007 41

TABLE 3-continued TABLE 3-continued Inhibition of MMP-3 by Plant Extracts Inhibition of MMP-3 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Veronica officinalis 27.0 Cucumis sativus Fanfare 35.4 Vicia faba 31.2 Cucurbita moschata 3O4 Vicia faba 44.7 Cucurbita pepo 23.8 Vigna angularia 40.8 Cucurbita pepo 46.6 Vigna angularis 39.4 Cuminum cyminum 23.1 Vignatinguiculata 26.1 Curcuna Zedoaria 20.8 Vitis sp. 624 Cymbopogon citratus 39.7 Vitis sp. 6.3.3 Doichus iabiab 25.8 Vitis sp. 82.O Dryopteris filix-mas S4.O Withania somnifera 22.4 Echinacea purpurea 20.4 Xanthium strumarium 20.7 Eriobotrya japonica 34.8 Zea mayS 26.1 Eriobotrya japonica 42.9 Zea mayS 67.5 Foericulum vulgare 33.1 Abies lasiocarpa 46.2 Fragaria X ananassa 20.3 Acorus calamus 21.8 Fragaria X ananassa 42.8 Actinidia arguita 64.6 Glycine max 26.3 Agropyron repens 48.3 Glycine max 3O.S Aichemilia nois 1OOO Gossypium herbaceum 22.5 Aichemilia nois 1OOO Guizotia abyssinica 46.6 Allium cepa 39.8 Hamamelis virginiana 33.1 Allium cepa 45.2 Hamamelis virginiana 33.1 Aium tuberosum 28.2 Hamamelis virginiana 44.8 Aium tuberosum 28.8 Hedeona pullegiodes 46.8 Alpinia officinarim 26.4 Helenium hoopesii 27.9 Amelianchiera initoia 78.3 Heianthus annus 22.7 Amelanchier Sanguinea X A. laevis 66.5 Heianthus strumosus 3O.O angelica archangelica 25.2 Heliotropium arborescens 53.7 Apium graveolens 43.3 Helleborus niger 4.O.S Aralia Cordata 31.5 Hibiscus cannabinus 34.0 Aralia nudicatiis 37.7 Hordeum vulgare subsp. Vulgare 1OOO Aralia nudicatiis 48.5 Huntilus lupulus 24.9 Aronia melanocarpa 26.0 Huntilus lupulus 55.1 Aronia melanocarpa 53.3 Huntilus lupulus 77.6 Aronia prunifolia 79.2 Huntilus lupulus 79.1 Artemisia absinthium 1OOO Huntilus lupulus 1OOO Artemisia dractinius 42.O Huntilus lupulus 1OOO Apertis esculenius 67.8 Huntilus lupulus 1OOO Beta vulgaris 27.9 Hypericum hennyi 1OOO Beta vulgaris 33.2 Hyperictim perforatin 99.3 Beta vulgaris 53.0 Hypomyces lactiflorum 2O.S Borago officinalis 55.7 Iris versicolor 48.5 Brassica Naptis 71.9 Juniperus communis 33.8 Brassica oleracea 37.0 Lactuca serioia 21.5 Brassica oleracea 46.9 Laportea Canadensis 37.7 Brassica rapa 36.7 Lavendula anguistifolia 91.7 Bromits inermis 42.8 Lepidium sativum 24.7 Calendula officinalis L. 28.4 Levisticum officinale 24.9 Cameliia Sinensis syn. Thea Sinensis 86.4 Lolium perenne 22.3 Capsicum annus 29.7 Lonicera ranosissina 42.5 Capsicum annus 43.7 Lonicera syringantha 21.1 Capsicum finitesCens (tabasco) 22.0 Maius 53.1 Carya cordiformis 27.5 Maius hupehensis (Pamp.) Rehd. 76.5 Chaerophyllum bulbosum 27.1 Maius sp. 39.8 Chaerophyllum bulbosum 1OOO Maius sp. 45.7 Chelidonium maius S4.O Maiva moschata 22.8 Chrysanthemum parthenium SO4 Malva Sylvestris 57.6 Chrysanthemum coronarium 25.8 Matteucia pensylvanica 20.1 Cichorium intybus 23.9 Melissa officinalis 55.0 Citritius ianatus 33.2 Mentha piperita 35.5 Citrulius lanatus (Garden baby) 21.4 Mentha piperita 43.9 Citrus innettoides 39.2 Mentha piperita 56.6 Citrus innon 60.4 Mentha pullegitim 33.3 Corchorus oitorius 28.6 Mentha pullegitim 56.2 Cornus Canadensis L. SO.O Mentha spicaia 43.4 Cornus Canadensis L. 80.6 Mentha spicaia 58.0 Cosmos Sulphureus 2O.S Nicotiana tabacum 27.3 Cosmos Sulphureus 27.0 Nigella Saiiva 25.1 Crataegus sp 43.9 Ocimum Basilicum 2O2 Crataegus submolis 24.2 Ocnothera biennis 37.8 Crataegus submolis 55.1 Origantin nationara 45.2 Cuctimis anguria 33.2 Origantim vulgare 21.3 US 2007/01 22492 A1 May 31, 2007 42

TABLE 3-continued TABLE 3-continued Inhibition of MMP-3 by Plant Extracts Inhibition of MMP-3 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Origantim vulgare 23.3 Tragopogon porrifolium R 29.8 Origantim vulgare 23.6 Tragopogon porrifolium O 58.0 Origantim vulgare 37.2 Triticale sp. O 25.3 Panicum miliaceum 20.6 Tropaeoluim mails O 46.9 Panicum miliaceum 30.7 Tropaeoluim mails O 55.8 Pastinaca Saiva 26.1 Tropaeoluim mails R 64.7 Pastinaca Saiiva 1OOO Tsiiga canOadensis R 39.2 Peucedanum of easeinum 39.6 Vaccinium anglisiifolium R 28.0 Peucedanum of easeinum 53.4 Vaccinium anglisiifolium S 29.6 Phaseolus vulgaris 21.8 Vaccinium anglisiifolium R 33.3 Phaseolus vulgaris 23.6 Vaccinium anglisiifolium Ait. R 1OOO Phaseolus vulgaris 59.8 Vaccinium macrocarpon S 25.1 Physalis alkekengi 55.5 Vaccinium macrocarpon R 27.4 Physalis pruinosa 24.8 Vaccinium macrocarpon O 35.4 Plantago major 77.1 Vaccinium macrocarpon R 8O.S Poa compressa 544 Vaccinium macrocarpon O 90.5 Polygonium chinense 36.3 Valeriana oficinalis O 33.0 Polygonium chinense 61.4 Veratrum viride S 46.8 Polygontin persicaria 21.3 Verbascum thapsus O 33.4 Populus incrassata 50.7 Vicia faba R 26.6 Populus incrassata 50.7 Vicia faba O 35.8 Populius X petrovskyana 66.7 Vigna angularia S 29.3 Prunus cerasifera 26.1 Vigna angularia O S4.O Prunus cerasifera 642 Vigna Sesquipedalis O 1OOO Psidium guaiaba 22.9 Vignatinguiculata S 49.5 Piaridium aquilinus 43.O Vitia sp. O 99.6 Pyrus pyrifolia 28.2 Vitis sp R SO.9 Rahmnus fiangula 25.9 Vitis sp. R 75.8 Raphantis Saivais 21.4 Weigela coracensis S 22.8 Raphantis Saivais 36.9 Weigela coracensis S 22.8 Rhamnus fianglia 43.2 Weigela hortensis R S4.9 Rheum rhabarbarium 28.5 Zea mayS O 74.3 Rheum x cuitorum 28.2 Rianits communis 32.4 Ribes nidigrolaria 28.5 Ribes nigrum 49.9 Rosa rugosa 29.1 0247 Rosmarinum officinalis 48.2 Rubus arcticus 59.1 TABLE 4 Rubus idealis 21.5 Rubus pubescens 51.8 Inhibition of MMP-9 by Plant Extracts Rubus thibetanus 33.7 Rumex patientia 34.4 Inhibition Rita graveolens 24.3 Latin name Stress Extract (%) Salvia (elegens) 37.2 Salvia (elegens) 42.9 Abelnochtis escientiis A. S 26.8 Salvia officinalis 67.3 Achillea millefolium A. S 41.6 Sambucus Canadensis 30.2 Aconium napeius A. O 47.7 Sanguisorba minor 21.0 Acorus calamus A. O 83.2 Sanguisorba minor 29.9 Actinidia arguita A. S 26.8 Sanguisorba minor 30.8 Adiantum pedatum A. O 20.7 Sanguisorba minor 445 Agastache foeniculum A. S 1OO.O Santoina 43.8 Agrimonia eupatoria A. W 21.4 Sarratuia tinctoria 37.7 Agropyron cristainin A. R 51.4 Satureia montana 4S.O Agropyron repens A. S 27.3 Satureia repandra 46.3 Agrostis alba A. R 40.6 Scorzorera hipanica 25.7 Agrostis Stofonifera A. R 35.4 Scuttellaria laterifiora 41.2 Alcea rosea A. S 45.8 Setaria itaica 33.4 Aikanna tinctoria A. S 42.5 Solidago Canadensis 78.5 Allium cepa A. O 49.7 Stachys affinis 1OOO Allium grande A. R 71.4 Stachys byzantina 1OOO Allium porrum A. S 28.0 Steiaria media (linne) Cyrillo 51.2 Allium porrum A. O 82.O Tanacetum vulgare 3O.S Aium sativum A. S 23.7 Teparty 31.7 Allium Schoenoprastin A. O 45.5 Teparty 39.7 Aium tuberosum A. V 20.1 Thymus serpyllum 29.9 Aium Tiberosum A. O 91.5 Thymus serpyllum 32.8 Althaea officinalis A. S 29.6 Thymus X citriiodorus 22.1 Amaranthus gangeticus A. O 25.1 Tiarella cordifolia 46.8 Amaranthus gangeticus A. R 31.1 Tragopogon porrifolium 26.3 Amaranthus gangeticus A. S 73.2 US 2007/01 22492 A1 May 31, 2007 43

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Amaranthus retroflexus 20.4 Chrysanthemum leucanthemum O 20.6 Ambrosia artemisiifolia SO.1 Chrysanthemum leucanthemum R 30.9 Amelanchier Sanguinea 37.6 Chrysanthemun coronarium (Chp R 26.4 Anthemis nobiis 40.4 Suey) Anthemis nobiis 66.7 Chrysanthemum coronarium 66.6 Anthemistinctorium 30.3 Cichorium intybus 44.7 Apium graveolens 71.2 Citritius ianatus 62.1 Arachis hypogaea 23.5 Citritius ianatus 70.6 Aralia Cordata 21.2 Corontis Canadensis 48.5 Aralia Cordata 56.3 Cosmos Sulphureus 23.4 Arctii in minus 31.1 Cosmos Sulphureus 37.0 Arctostaphylos tiva-ursi 31.2 Crataegus sp 32.4 Arctostaphylos tiva-ursi 31.2 Crataegus sp 45.5 Arctostaphylos tiva-ursi 59.7 Crataegus sp 1OO.O Armoracia rusticana 25.1 Crataegus submolis 45.5 Armoracia rusticana 56.2 Cryptotaenia Canadensis 26.4 Aronia melanocarpa 26.8 Clictimis Anguria 27.2 Aronia melanocarpa 41.3 Clictimis anguria 36.6 Aronia melanocarpa 44.8 Clictimis anguria 38.5 Aronia melanocarpa 47.7 Cucumis meio 59.2 Aronia melanocarpa 55.7 Ciclinis Saiivus 39.8 Aronia melanocarpa 1OO.O Ciclinis Saiivus 49.4 Arrhenaiherum eiatius 40.4 Ciclinis Saiivus 54.4 Artemisia dractinctius S1.1 Cucurbita Maxina 46.7 Asparagus officinalis 20.9 Cucurbita moschata 32.1 Asparagus officinalis 32.6 Cucurbita pepo 37.0 Aster sp 29.5 Curburbita pepo 410 Aster sp 80.0 Curburbita pepo 43.9 Atropa belladonna 47.4 Curcuna Zedoaria 67.6 Beta vulgaris 25.3 Curcurbita maxima 25.8 Beta vulgaris 26.6 Cymbopogon citratus 26.7 Beta vulgaris 34.O Dactylis glomerata 27.2 Beta vulgaris 42.O Datisca Cannabina 26.9 Beta vulgaris 44.0 Datisca Cannabina 38.0 Beta vulgaris spp. Maritima 44.0 Daiiciis Caroia 30.8 Beta vulgaris var. Condiwata 35.4 Daiiciis Caroia 31.9 Brassica naptis 24.6 Dirca palustris 27.3 Brassica naptis 53.1 Dirca palustris 34.2 Brassica naptis 1OO.O Doicos Labiah 22.O Brassica nigra 24.2 Doicos Labiah 25.3 Brassica oleracea 33.0 Dryopteris filix-mas 24.9 Brassica oleracea 36.0 Dryopteris filix-mas 40.6 Brassica oleracea 36.2 Eleusine Coracana 2O2 Brassica oleracea 73.1 Eleusine Coracana 20.9 Brassica Oieracea 1OO.O Eleusine Coracana 71.1 Brassica rapa 31.0 Elymus incetts 45.4 Brassica rapa 38.6 Erigeron Canadensis 35.7 Brassica rapa 42.8 Erica vesicaria 59.9 Brassica rapa 48.8 Fagopyrim esculenium 20.7 Brassica rapa 68.2 Fagopyri in tartarictim 30.3 Brassica rapa 89.2 Fagopyri in tartarictim 33.2 Bromits inermis 51.4 Festica rubra 31.8 Campaniiia raptinctiitis 25.1 Foeniculum Vulgare 27.4 Canna editiis 31.1 Foeniculum vulgare SO.6 Canna editiis 47.6 Forsythia intermedia 1OO.O Canna editiis 68.9 Fragaria X ananassa 3O.O Capsella bursa-pastoris 32.5 Fragaria X ananassa 36.3 Capsicum annittin 22.O Gaium odoratum 26.9 Capsicum annittin 24.O Gaultheria hispidula 28.4 capsicum annuitin 55.7 Gaultheria hispidula 40.7 Capsicum finitesCens 30.3 Gentiana littea 34.7 Capsicum finitesCens 34.7 Giechona hederacea 37.6 Carthamus tinctorius 28.5 Glycine max 38.1 Cartin carvi 38.6 Glycine Max 56.4 Chelidonium maius 27.9 Glycine max 71.4 Chenopodium bonus-henricus 47.4 Glycyrrhiza glabra 62.6 Chenopodium bonus-henricus 20.7 Glycyrrhiza glabra 1OO.O Chenopodium bonus-henricus 23.2 Guizotia abyssinica 91.9 Chenopodium bonus-henrict is 62.8 Hamamelis virginiana 410 Chenopodium quinoa 23.1 Hamamelis virginiana 74.6 Chenopodium quinoa 34.7 Hedeona pullegioides 22.O US 2007/01 22492 A1 May 31, 2007 44

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Heianthus tuberosus 21.2 Physalis alkekengi 31.6 Heianthus tuberosus 51.5 Physalis ixocarpa 45.2 Helichrysum angustifolium 21.O Physalis Ixocarpa 65.3 Heliotropium arborescens 54.1 Physalis Pruinosa 87.3 Helleborus niger 37.8 Phytolacca americana 49.6 Hordeum hexastichon 38.0 Phytolacca americana 89.8 Hyssopus officinalis 25.1 Pimpinella anisum 1OO.O Initia heienium 29.7 Plantago Coronopus 48.3 Isais inctoria 41.5 Plantago Coronopus 89.3 Lactuca Serrilla 41.3 Plantago major 21.8 Lactuca serioia 46.6 Poa compressa 22.4 Laportea Canadensis 26.3 Poa compressa 49.3 Lathyrus Saivais 22.2 Poa pratensis 22.4 Lathyrus Saivais SO.2 Polygontin pensylvanictim 43.3 Lathyrus Sylvestris 31.3 Polygontin persicaria 21.6 Lathyrus Sylvestris 31.8 Polygontin persicaria 38.5 Latiri is nobilis 25.7 Potentiiia anserina 26.3 Latiri is nobilis 3O.O Potentiiia anserina 31.2 Lavandhiia iaiifolia 40.3 Poterium Sanquisorba 29.2 Leonirits cardiaca 27.0 Pteridium aquilinum 27.3 Lepidium sativum 4.1.8 Raphantis Saivais 22.7 Levisticum officinale 29.0 Raphantis Saivais 30.8 Levisticum officinale 44.9 Raphantis Saivais 40.2 Linaria vulgaris milier 23.6 Raphantis Saivais 71.5 Lintin itsitatissini in 33.3 Raphantis Saivais 1OO.O Lolium multiflorum 29.0 Rheum rhabarbarium 21.3 Lolium perenne S2.O Rheum rhabarbarium 67.9 Lotus corniculatus 62.9 Rheum rhabarbarium 72.4 Lotus tetragonolobits 62.9 Ribes nidigrolaria 32.6 Lycopersicon esculenium 26.1 Ribes nidigrolaria 64.6 Lycopersicon esculenium 33.0 Ribes nigrum 23.6 Maiva moschata 31.8 Ribes nigrum 27.2 Malva Sylvestris 21.4 Ribes nigrum 410 Maiva verticiliata 43.4 Ribes nigrum 65.8 Matteucia pensylvanica 26.9 Ribes Nigrum 1OO.O Medicago Saiiva 20.4 Ribes Saivum 75.4 Meilotus albus 53.9 Ribes Sylvestre 27.7 Melissa officinalis 21.4 Ribes Sylvestre 1OO.O Melissa officinalis 36.8 ribes tiva-Crispa 24.4 Melissa officinalis 53.7 Ribes Uva-crispa 36.6 Mentha piperita 57.7 Ricinits communis 21.6 Mentha pullegitim 66.1 Rosa rugosa 30.6 Mentha spicaia 67.7 Rosa rugosa 36.2 Mentha suaveolens 51.8 Rosa rugosa 39.3 Monoraica charantia 29.7 Rosmarinus officinalis 27.2 Monoraica charantia 72.1 Rosmarinus officinalis 45.7 Nicotiana rustica 30.3 Rubus allegheniensis 53.7 Nicotiana rustica 59.1 Rubus Canadensis 27.0 Nicotiana tabacum 39.0 Rubus Canadensis 410 Nicotiana tabacum 47.6 Rubus Canadensis 41.2 Nicotiana tabacum 1OO.O Rubus Canadensis 45.1 Nigella Saiiva 59.4 Rubus idaeus 24.3 Oenothera biennis 21.3 Rubus idaeus 39.7 Oenothera biennis 36.7 Rubus idaeus 62.2 Origantim vulgare 21.3 Rubus idaeus 37.0 Origantim vulgare 42.7 Rumex acetoseiia 75.8 Oryza sativa 56.5 Rumex acoiosa 25.5 Oxyria digyna 35.1 Rumex crispiis 73.3 Oxyria digyna 76.4 Rumex crispiis 6O.S Pastinaca Saiiva 20.3 Rumex patientia 49.4 Pastinaca Saiiva 23.2 Rumex patientia 65.8 Pastinaca Saiiva 42.1 Rumex Scittatus 25.5 Pastinaca Saiiva 46.9 Rumex Scittatus 61.9 Phaiaris canariensis 20.3 Rumex Scittatus 93.8 Phaiaris canariensis 8O.S Rita graveolens 25.8 Phaseolus mingo 51.3 Rita graveolens 27.1 Phaseolus mingo 74.1 Salix purpurea 22.1 Phaseolus vulgaris 23.0 Salix purpurea 33.8 Phaseolus vulgaris 51.4 Salvia elegans 23.7 Phaseolus vulgaris 62.6 Salvia officinalis 20.8 Phlox paniculata 410 Salvia officinalis 31.4 US 2007/01 22492 A1 May 31, 2007 45

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Saivia Sciarea A. S 28.0 Adiantum pedatum G R 31.6 Satureia montana A. W 21.7 Adiantum pedatum G S 31.7 Scuttellaria latenflora A. S 54.1 Ageratum conyzoides G S 23.1 Secale cereale A. V 22.6 Agropyron cristainin G R 64.1 Secale cereale A. S 22.9 Agropyron repens G S 29.2 Secale cereale A. W 26.9 Agropyron repens G O 32.6 Sesamun indicum A. O 21.2 Agrostis Stoionifera G R 34.4 Setaria itaica A. O 27.0 Alcea rosea G S 22.7 Siim Sisartin A. R 32.6 Aichemia nois G S 3O.S Siim Sisartin A. O 42.7 Aichemia nois G W 33.2 Soianum duticamara A. S 43.3 Allium ampeioprastin G O 53.4 Soianum duticamara A. O 48.6 Allium cepa G S 22.5 Soianum melanocerastin A. O 21.3 Allium cepa G O 60.7 Soianum melongena A. R 2O.S Allium Schoenoprastin G S 21.1 Soianum melongena A. V 35.6 Allium Schoenoprastin G O 60.4 Soianum melongena A. O 49.4 Aium tuberosum G S 38.8 Soianum melongena A. S 65.2 Aium tuberosum G O 74.4 Solidago sp A. R 32.7 Althaea officiana is G S S4.9 Spinacia oleracea A. S 410 Amaranthus candaihats G O 42.6 Stachys affinis A. R 22.5 Amaranthus caudathus G W 27.1 Stachys affinis A. S 43.9 Amaranthus gangeticus G S 56.8 Stachys affinis A. O 92.0 Amaranthus gangeticus G S 74.4 Symphytum officinale A. S 28.0 Ambrosia artemisiifolia G R 49.0 Tanacetum cinerariifolium A. O 20.3 Amelanchier Sanguinea G W 45.2 Tanacetum cinerariifolium A. R 69.7 Angelica archangelica G S 20.9 Tanacetum vulgare A. O 2O2 Anthemis nobiis G R S8.9 Tanacetum vulgare A. S 84.2 Apium graveolens G O 30.4 Teucrium chamaedrys A. O 20.4 Apium graveolens G S 36.4 Teucrium chamaedrys A. R 20.4 Apium graveolens G R 60.6 Thymus serpyllum A. W 24.3 Arachis hypogaea G W 26.O Thymus vulgaris A. S 42.5 Aralia Cordata G S 66.O Thymus X citriiodorus A. W 27.4 Arctii in minus G O 26.6 Tragopogon porrifolius A. W 21.9 Arctii in minus G R 30.8 Tragopogon porrifolius A. V 26.2 Arctostaphylos tiva-ursi G S 29.3 Trifolium hybridum A. R 30.9 Arctostaphylos tiva-ursi G O 38.8 Trifolium pannonicum A. R 410 Arctostaphylos tiva-ursi G R 80.2 Trifolium repens A. R 51.3 Armoracia rusticana G S 62.7 Trigonelia foenim graecum A. S 44.2 Aronia melanocarpa G O 26.7 Triticum spelta A. S 3O.O Aronia melanocarpa G V 1OO.O Triticum turgidium A. S 31.3 Aronia melanocarpa G R 1OO.O Typha latifolia A. S 57.7 Aronia melanocarpa (Michx.) Ell. G W 39.1 Urtica dioica A. O 26.5 Artemisia dractinctius G O 44.3 Urtica dioica A. S SO.2 Artemisia dractinctius G S 65.4 Vaccinium Corymbosum A. W 39.9 Asclepias incarnata G R 20.3 Vaccinium Corymbosum A. S 64.8 Asparagus officinalis G O 22.3 Vaccini in augustifolium A. R 44.8 Asparagus officinalis G S 26.6 Vaccinum macrocarpon A. S 1OO.O Asparagus officinalis G W 28.7 Veratrum viride A. S 29.1 Aster sp G O 34.3 Veratrum viride A. O 31.8 Aster sp G R 62.6 Verbascum thapsus A. S 42.6 Atropa belladonna G S 34.9 Verbascum thapsus A. O 75.2 Beta vulgaris G R 28.3 Viburnum trilobium A. V 97.4 Beta vulgaris G R 42.2 Vicia Saiva A. R 53.3 Beta vulgaris G O 47.0 Vicia viliosa A. R 48.9 Beta vulgaris spp. Maritima G O 46.7 Vignatinguiculata A. R 27.0 Brassica cepicepa G R 26.7 Vignatinguiculata A. O 44.8 Brassica cepicepa G S 68.3 Vignatinguiculata A. S 55.5 Brassica iuncea G O 45.0 Vinca minor A. S 35.1 Brassica iuncea G S 66.1 Vitis sp. A. V 52.2 Brassica Naptis G S 27.5 Vitis sp. A. S 59.6 Brassica Naptis G R 37.6 Vitis sp. A. R 87.8 Brassica naptis G O 94.8 Xanthium Sibiricum A. S 57.1 Brassica nigra G S 36.4 Zea mayS A. V 26.1 Brassica oleracea G R 38.7 Zea mayS A. W 32.1 Brassica oleracea G W 39.0 Zea Mays A. O 38.7 Brassica oleracea G R 49.4 Achillea millefolium G S 45.5 Brassica oleracea G S 76.1 Aconium napeius G S 24.O Brassica oleracea G O 1OO.O Aconium napeius G O 53.9 Brassica rapa G R 21.1 Acorus calamus G O 87.6 Brassica rapa G S 64.O Acorus calamus G S 1OO.O Brassica rapa G O 1OO.O Actinidia arguita G S 33.8 Bromits inermis G R 36.7 US 2007/01 22492 A1 May 31, 2007 46

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Campaniiia raptinctiitis 59.9 Gaium odoratum 32.4 Canna editiis 20.8 Gaium odoratum 1OO.O Canna editiis 83.1 Gaultheria hispidula 48.4 Capsicum annittin 2O2 Gaultheria hispidula 80.4 Capsicum annittin 29.6 Gaultheria hispidula 1OO.O Capsicum annittin 51.5 Gaultheria procumbens 26.9 Capsicum annittin 60.8 Gaultheria procumbens S4.3 Capsicum finitesCens 32.8 Giechona hederacea 26.6 Carthamus tinctorius 29.8 Glycine max 52.5 Cartin carvi 30.4 Glycine max 67.9 Chelidonium maius 39.9 Glycine max 75.8 Chenopodium bonus-henricus 63.0 Glycyrrhiza glabra 21.4 Chenopodium quinoa 34.1 Glycyrrhiza glabra 21.6 Chenopodium quinoa 42.8 Glycyrrhiza glabra 1OO.O Chenopodium quinoa 46.1 Guizotia abyssinica 91.4 Chichorium endivia subspendivia 22.O Hamamelis virginiana 39.8 Chichorium endivia subspendivia 22.9 Hamamelis virginiana 78.8 Chrysanthemum coronarium 23.2 Hamamelis virginiana 96.6 Chrysanthemum coronarium 68.4 Hedeona pullegioides 45.4 Chrysanthemum leucanthemum 2O.S Helenium hoopesii 22.6 Cicer arrieintin 25.7 Helenium hoopesii 52.8 Cichorium intybus S1.1 Helianthus anniitis 22.O Cichorium intybus 53.4 Helianthus anniitis 31.6 Citritius ianatus 36.5 Heianthus strumosus 3O.S Citritius ianatus 71.5 Heianthus strumosus 71.7 Coix Lacryma-Jobi 21.O Heianthus tuberosus 21.2 Cornus Canadensis 34.8 Heianthus tuberosus 50.7 Crataegus sp S4O Heianthus tuberosus L. 24.9 Crataegus submolis 31.3 Heliotropium arborescens 40.O Cryptotaenia Canadensis 32.1 Heliotropium arborescens 45.6 Cuctimis anguria 27.3 Helleborus niger 38.0 Cuctimis anguria 32.5 Hordeum vulgare 21.5 Ciclinis Saiivus 39.4 Huntilus lupulus 35.1 Ciclinis Saiivus 69.4 Hyperictim sp 26.1 Cucurbita maxima 34.1 Hyssopus officinalis 74.5 Cucurbita maxima 42.6 Iberis amara 20.9 Cucurbita moschata 32.O Iberis amara 21.7 Cucurbita moschata 39.2 Initia heienium 27.6 Cucurbita pepo 28.8 Ipomoea batatas 37.5 Cucurbita pepo 32.6 Isais inctoria 48.0 Curcuna Zedoaria 23.3 Lachica Serroia 53.0 Curcuna Zedoaria 57.6 Lactica Saiiva 24.5 Cymbopogon citratus 70.1 Laportea Canadensis 36.0 Cynara Scolymits 2O2 Laportea Canadensis 81.7 Cynara Scolymits 37.5 Lathyrus sativus 37.8 Cynara Scolymits 88.7 Lathyrus Sylvestris 40.7 Cypertis esculenius 66.7 Lathyrus Sylvestris 79.1 Datura metel 29.2 Latiri is nobilis 22.7 Daitira Stranonium 27.6 Lavandhiia angustifolia 31.7 Daiiciis Caroia 24.2 Lavandhiia iaiifolia 27.2 Daiiciis Caroia 29.3 Ledum groenlandictim 61.1 Dipsacus sativus 48.7 Leonirits cardiaca 22.6 Dirca palustris 29.9 Lepidium sativum 23.3 Dirca palustris 36.4 Levisticum officinale 23.1 Doichos Labiah 35.8 Levisticum officinale 27.5 Doichos Labiah 74.5 Levisticum officinale 41.3 Dryopteris filix-mas 27.9 Lintin itsitatissini in 21.4 Dryopteris filix-mas 42.6 Lolium perenne 32.7 Echinochloa frumentacea 68.4 Lotus corniculatus 54.2 Eleusine Coracana 47.8 Maius hupehensis 26.4 Elymus it incetts 42.7 Maiva verticiliata 37.9 Erigeron Canadensis 37.8 Matricaria recuita S.O.3 Erigeron speciosus 34.6 Medicago Saiiva 29.1 Errhenaiherum eiatius 34.4 Meilotus albus S2.1 Fagopyri in tariarictim 31.4 Melissa officinalis 22.7 Foeniculum vulgare 28.0 Melissa officinalis 35.9 Foeniculum vulgare 44.6 Melissa officinalis 38.6 Foeniculum vulgare 68.9 Mentha piperita 64.4 Foeniculum Vulgare 1OO.O Mentha suaveoiens 22.5 Forsythia intermedia 1OO.O Monoraica charantia 29.3 Forsythia X intermedia 79.5 Monoraica charantia 90.6 US 2007/01 22492 A1 May 31, 2007 47

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Nepeta cataria G R 50.5 Rubus idealis G O 32.5 Nicotiana rustica G O 35.3 Rubus idealis G S 47.0 Nicotiana rustica G S 1OO.O Rubus occidentais G S 39.4 Nicotiana tabacum G S 31.6 Rubus occidentais G R 74.1 Nicotiana tabacum G O 1OO.O Rumex acetosa G W 45.6 Nigella Saiiva G R 24.2 Rumex acetoseiia G W 22.8 Ocimum basicum G S 30.6 Rumex acetoseiia G V 31.5 Oenothera biennis G O 48.0 Rumex crispiis G O 25.9 Oenothera biennis G R 76.6 Rumex crispiis G R 70.3 Origantim vulgare G V 41.3 Rumex patientia G O 39.8 Oryza Saiva G O 22.1 Rumex patientia G S 54.2 Oxyria digyna G O 26.5 Rumex Scittatus G W 23.8 Oxyria digyna G V 70.3 Rumex Scittatus G V 69.9 Panicum miliaceum G O 94.4 Rumex Scittatus G O 78.8 Pastinaca Saiiva G R 29.4 Rita graveolens G R 30.7 Pastinaca Saiiva G S 79.2 Rita graveolens G S 61.5 Pennisetum alopectiroides G O 22.O Salvia elagens G W 25.4 Petasites japonicus G S 29.2 Salvia elegans G S 31.1 Peucedanum of easeinum G O 21.3 Sambucus Canadensis G W 80.6 Phacelia tanacetifolia G R 23.5 Sambucus ebulus G W 26.1 Phaiaris arundinacea G R 47.5 Sambucus ebulus G V 34.4 Phaiaris canariensis G R 23.1 Sambucus ebulus G S 37.8 Phaiaris canariensis G O 1OO.O Sanguisorba officinalis G R 1OO.O Phaseolus coccineus G O 37.0 Santolina Chamaecyparissus G R 21.7 Phaseolus coccineus G R 74.1 Santolina Chamaecyparissus G S 25.2 Phaseolus mingo G O 42.2 Satureia montana G O 21.2 Phaseolus mingo G S 52.2 Scuttellaria laterifiora G S 37.0 Phaseolus vulgaris G V 35.5 Secale cereale G S 26.7 Phaseolus vulgaris G S 48.0 Secale cereale G W 27.3 Phaseolus vulgaris G O S8.1 Serrainia tinctoria G S 36.2 Phlox paniculata G S 32.2 Serrainia tinctoria G O 70.3 Phlox paniculata G O 40.1 Sesamun indicum G O 27.6 Physalis ixocarpa G O 20.6 Sesamun indicum G S 44.3 Physalis pruinosa G O 80.0 Silybum marianum G S 34.7 Phytolacca americana G S 62.O Siim Sisartin G O 79.0 Phytolacca americana G O 1OO.O Soianum duticamara G R 25.2 Pimpinella anisum G S 37.3 Soianum duticamara G S 64.6 Pistin Saivitin G W 34.4 Soianum melongena G S 36.6 Pistin Saivitin G O 6.3.3 Soianum melongena G O 40.1 Plantago Coronopus G O 42.7 Soianum melongena G V SO.O Plantago Coronopus G S 46.4 Soianum melongena G S 74.9 Plantago major G O 28.3 Soianum tuberosum G S 39.1 Plantago major G S 41.4 Soianum tuberosum G O 39.2 Plectranthus sp. G S 29.3 Solidago sp G R 30.7 Poa compressa G R 22.1 Sorghum cafiorum G O 87.9 Poa compressa G S 45.5 Sorghum dochna G W 20.6 Poa praiensis G R 35.7 Sorghum dochna G O 20.6 Polygontin pensylvanictim G S 38.3 Sorghum dochna G S 34.1 Polygontin persicaria G S 31.0 Sorghum dochna G O 97.0 Potentiiia anserina G O 46.8 Sorghum durra G O 30.6 Poterium sanquisorba G S 24.7 Sorghum durra G S 30.6 Poterium sanquisorba G W 30.6 Sorghum durra G O 48.6 Prunus cerasifera G R 45.9 Sorghum Sudanense G S 21.7 Pteridium aquilinum G S 22.4 Sorghum Sudanense G O 24.6 Raphantis Raphanistrain G S 36.5 Sorghum Sudanense G V 32.1 Raphantis Raphanistrain G O 75.O Spinacia oleracea G S 53.2 Raphantis Saivais G R 20.8 Stachys Affinis G S 2S.O Raphantis Saivais G R 27.5 Stachys Affinis G R 27.8 Raphantis Saivais G S 35.4 Stachys Affinis G O 1OO.O Rheum rhabarbarium G S 27.0 Symphytum officinale G W 21.7 Ribes Grossuiaria G W 33.7 Symphytum officinale G O 25.2 Ribes nidigrolaria G S 30.7 Symphytum officinale G S 34.6 Ribes nidigrolaria G V 40.5 Tanacetum cinerariifolium G R 52.4 Ribes nigrum G V 35.9 Tanacetum vulgare G R 27.1 Ribes nigrum G W 58.6 Tanacetum vulgare G S 72.7 Ribes Silvestris G V 26.9 Teucrium chamaedrys G R 24.6 Ribes Silvestris G W 1OO.O Teucrium chamaedrys G O 52.8 Ricinits communis G R 21.8 Thymus fragantissuinus G R 1OO.O Rosmarinus officinalis G S 24.7 Thymus vulgaris G V 24.2 Rosmarinus officinalis G W 30.9 Thymus X citriiodorus G S 23.7 Rosmarinus officinalis G R 60.3 Tiarella cordifolia G S 20.8 US 2007/01 22492 A1 May 31, 2007 48

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Tiarella cordifolia 30.8 Allium grande 60.4 Tragopogon porrifolius 22.8 Allium Schoenoporastin 65.8 Trifolium hybridum 24.7 Allium Schoenoprastin 31.0 Trifolium pannonicum 65.5 Aium tuberosum 22.8 Trifolium repens 57.5 Aium tuberosum 99.7 Trigonelia foeningraeciin 37.6 Althaea officiana is 22.8 Triticum fungidum 56.5 Althaea officinalis 22.1 Triticum spelta 40.8 Amaranthus candaihats 43.9 Tropaeoluim maints 76.1 Amaranthus gangeticus 30.3 Typha latifolia 43.3 Amaranthus gangeticus 66.O Urtica dioica 40.3 Ambrosia artemisiifolia 58.7 Vaccinium anglisiifolium 42.4 Amelianchiera initoia 70.5 Vaccinium corymbosum 61.5 Amelanchier Sanguinea 37.3 Vaccinium macrocarpon 43.7 Ananas Conostis 23.8 Vaccini in angustifolium 23.1 Ananas Conostis 95.0 Veratrum viride 43.6 Ananas Conostis 99.6 Verbascum thapsus 37.8 angelica archangelica 3O.S Verbascum thapsus 87.O angelica archangelica 38.9 Veronica officinalis 3O.S Anthemis nobiis 41.4 Viburnum trilobium 49.4 Anthemis nobiis 72.8 Viburnum trilobium 1OO.O Anthemistinctorium 27.3 Viburnum trilobium 1OO.O Anthriscus cerefolium 35.8 Vicia faba 50.5 Apium graveolens 31.7 Vicia Saiva 42.4 Apium graveolens 32.4 Vicia viliosa 89.2 Apium graveolens 56.6 Vigna angularia 28.1 Aralia Cordata 29.2 Vigna angularia 71.5 Aralia Cordata 45.0 Vigna unguiculata 21.O Arctii in minus 25.8 Vignatinguiculata 38.7 Arctostaphylos tiva-ursi 31.0 Vignatinguiculata 61.1 Arctostaphylos tiva-ursi 35.2 Vinca minor 33.6 Arctostaphylos tiva-ursi 58.6 Vinca minor 34.3 Armoracia rusticana 24.9 Vitis sp. 29.0 Armoracia rusticana S2.9 Vitis sp. SO.2 Aronia melanocarpa 40.O Vitis sp. 53.3 Aronia melanocarpa 91.9 Vitis sp. 63.0 Aronia prunifolia 1OO.O Vitis sp. 86.6 Arrhenatherum elatius 22.8 Withania somnifera 20.3 Artemisia draculus 74.9 Xanthium Sibiricum 34.7 Artemisia dractinctius 47.8 Xanthium strumarium 23.2 Asclepias incarnata 2O.S Zea mayS 20.1 Asciinidia chinensis 43.4 Zea mayS 45.9 Asciinidia chinensis 66.4 Zea mayS 97.5 Asparagus officinalis 91.3 Abelnochtis escientiis 24.8 Asparagus officialis 23.3 Abies lasiocarpa 44.7 Asparagus officialis 44.7 Achillea millefolium 24.1 Aster Linne 47.5 Achillea millefolium 59.2 Aster sp 62.O Aconium napeius 40.6 Atriplex hortensis 54.6 Aconium napeius 41.6 Atropa belladonna 20.1 Acorus calamus 47.1 Atropa belladonna S1.O Actinidia arguita 21.8 Avena Saiiva 24.8 Adiantum pedatum 26.8 Avena Saiiva 26.4 Adiantum pedatum 45.8 Averrhoa caramboia 23.4 Adiantum pedatum 86.0 Apertis esculenius 46.2 Agarict is bisports 26.3 Beta vulgaris 28.2 Agarict is bisports 29.8 Beta vulgaris 30.4 Agarict is bisports 36.9 Beta vulgaris 56.8 Agarict is bisports 44.0 Beta vulgaris spp. Maritima 23.6 Agarict is bisports 46.0 Betula glandulosa 22.2 Agastache foeniculum 70.O Betula glandulosa 22.2 Ageratum Conyzoides 31.7 Betula glandulosa 25.7 Agropyron cristainin 86.9 Betula glandulosa 32.9 Agropyron repens 49.6 Boietus eduis 36.2 Agrostis alba 21.9 Boietus eduis 90.2 Agrostis Stoionifera 35.8 Borago officinalis 27.9 Alcea rosea 35.2 Borago officinalis 76.1 Aichemilia nois 37.9 Brassica cepicepa 65.4 Allium ampeioprastin 48.0 Brassica cepicepa 71.5 Allium ascalonictim 26.2 Brassica Chineusis 27.1 Allium ascalonictim 77.2 Brassica iuncea S1.O Allium cepa 92.6 Brassica iuncea 66.O US 2007/01 22492 A1 May 31, 2007 49

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Brassica iuncea 74.1 Crataegus sp 28.0 Brassica Naptis 22.O Crataegus sp 60.9 Brassica Naptis 34.O Crataegus submolis 25.5 Brassica Naptis 1OO.O Crithmin maritima SO.6 Brassica nigra 26.7 Cryptotaenia Canadensis 21.2 Brassica nigra 27.4 Cryptotaenia Canadensis 26.O Brassica nigra 82.5 Cryptotaenia Canadensis 40.O Brassica oleracea 21.2 Clictimis anguria 38.7 Brassica oleracea 22.1 Clictimis anguria 46.6 Brassica oleracea 26.2 Cucumis meio 30.3 Brassica oleracea 27.2 Cucumis meio 46.2 Brassica oleracea 31.3 Cucumis metuliferus 32.O Brassica oleracea 46.5 Cucumis sativus Fanfare 40.3 Brassica oleracea 71.2 Cucurbita maxima 23.6 Brassica oleracea 93.5 Cucurbita maxima 33.1 Brassica rapa 25.6 Cucurbita maxima 55.2 Brassica rapa 33.9 Cucurbita moschata 20.1 Brassica rapa 56.0 Cucurbita moschata 26.7 Brassica rapa 69.7 Cucurbita moschata 41.7 Brassica rapa 1OO.O Cucurbita pepo 41.9 Bromits inermis 57.3 Cucurbita pepo 82.9 Campaniiia raptinctiitis 77.5 Curcuna Zedoaria 1OO.O Canna editiis 75.6 Cydonia oblonga 42.9 Canthareilus ciparium 52.5 Cynara Scolymits S1.6 Capsella bursa-pastoris 35.9 Cynara Scolymits 60.9 Capsicum annus 43.9 Dactiis Giomerata 25.7 Capsicum annittin SO.1 Daitira Stranonium 21.9 Capsicum futescens 28.9 Daiiciis Caroia 25.9 Carica papaya 31.1 Dioscorea batatas 47.6 Carthamus tinctorius 37.3 Dioscorea batatas 83.1 Cartin carvi 30.1 Diospiros Kaki 34.9 Castanea spp. 21.7 Dirca palustris 27.6 Chaerophyllum bulbosum 46.0 Dirca palustris 90.4 Chamaemeium nobile 36.8 Doichus iabiab 66.4 Chamaemeium nobile 48.4 Doichus iabiab 85.3 Chelidonium maius 46.6 Dryopteris filix-mas 21.9 Chenapodium bonus-henricus 22.4 Dryopteris filix-mas 77.9 Chenopodium bonus-henricus 57.6 Echinacea purpurea 48.6 Chenopodium quinoa 35.5 Eleusine Coracana 45.2 Chenopodium quinoa 54.4 Elymus incetts 410 Chrysanthemum leucanthemum 26.5 Erigeron Canadensis 31.4 Chrysanthemun coronarium (Chp 48.4 Eriobotrya japonica 28.3 Suey) Erica vesicaria 44.9 Chrysanthemum coronarium 38.2 Fagopyrim esculenium 76.7 Chrysanthemum coronarium 63.9 Fagopyri in tartarictim 42.6 Cicer arrieintin 2O.O Festica rubra 29.6 Cichorium endivia 25.6 Festica rubra 42.9 Cichorium endivia crispa 38.4 Foeniculum vulgare 22.1 Cichorium intybus 30.2 Foericulum vulgare 21.6 Cinicifiiga racemosa 33.7 Foericulum vulgare 84.8 Citrulius colocynthus 20.4 Forsythia intermedia 70.8 Citritius ianatus 68.3 Forsythia X intermedia 60.2 Citritius ianatus 31.9 Fortunella spp 35.7 Citrus innettoides 20.4 Fortunella spp 50.7 Citrus innettoides 37.5 Fortunella spp 74.5 Citrus innon 47.7 Fragaria 24.8 Citrus innon 72.4 Fragaria 52.4 Citrus paradisi 23.8 Fragaria 1OO.O Citrus paradisi 33.4 Fragaria X ananassa 29.3 Citrus reticulata 20.4 Gaium odoratum 26.O Citrus reticulata 20.9 Gaultheria hispidula 40.3 Citrus reticulata 26.O Ginkgo biloba 27.0 Citrus reticulata 40.4 Ginkgo biloba 68.9 Citrus reticulata SO.O Giechona hederacea 20.4 Citrus reticulata 79.2 Giechona hederacea 30.4 Citrus Sinensis 25.3 Glycine max 26.6 Citrus Sinensis 59.8 Glycine max 47.4 Coix Lacryma-Jobi 2O.O Glycine max 82.O Corchorus oitorius 38.9 Glycyrrhiza glabra 35.4 Cornus Canadensis 35.6 Glycyrrhiza glabra 40.5 Cosmos Sulphureus 51.4 Glycyrrhiza glabra 1OO.O US 2007/01 22492 A1 May 31, 2007 50

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name (%) Latin name Ex 8. C t (%) Gossypium herbaceum 36.1 Maiva moschata 41.1 Guizotia abyssinica 28.9 Malva Sylvestris 36.4 Guizotia abyssinica 40.4 Malva Sylvestris 47.4 Hamamelis virginiana 52.4 Maiva verticiliata 42.7 Hamamelis virginiana 67.5 Mangifera indica 3O.S Hamamelis virginiana 84.1 Manihot esculenta syn. M. utilissima 38.3 Hedeona pullegiodes 57.4 Manihot esculenta syn. M. utilissima SO.4 Helenium hoopesii 33.7 Manihot esculenta syn. M. utilissima 86.5 Helenium hoopesii 49.0 Meilotus aiba 30.4 Heianthus annus 53.4 Melilotus officinalis 68.1 Heianthus strumosus 20.3 Melissa officinalis 33.7 Heianthus strumosus 71.7 Melissa officinalis 34.7 Heianthus tuberosa 22.8 mentha arvensis 53.7 Heianthus tuberosus L. 22.6 Mentha suaveoiens 26.8 Heianthus tuberosus L. 55.0 Menyanthes trifoliata 32.8 Helichrysum angustifolium 67.0 Miscanthus sinensis Andress 22.7 Heliotropium arborescens S8.9 Monoraica charantia 55.5 Helleborus niger 31.9 Monarda didyma 26.8 Hibiscus cannabinus 48.9 Monarda fistulosa 21.5 Hordeum vulgare 29.2 Montia perfoliata 26.6 Huntilus lupuits 22.4 Musa paradisiaca 29.0 Huntilus lupuits 39.1 nastairtium officinale 35.4 Huntilus lupuits 63.1 Nepeta cataria 26.5 Huntilus lupuits 1OO.O Nepeta cataria 27.5 Hydrastis Canadensis 2O2 Nepeta cataria 41.9 Hydrastis Canadensis 31.0 Nephelium longana out Euphoria 43.4 Hyoscyami is niger 56.8 longana Hypericum henryi 48.8 Nicotiana rustica 26.O Hyperictim perforatin 48.1 Nicotiana rustica 32.7 Hyperictim perforatin 63.7 Nicotiana tabacum 25.1 Hypomyces lactiflorum 44.8 Nicotiana tabacum 77.7 Hypomyces lactiflorum 60.9 Nigella Saiiva 59.3 Hyssops officinalis 22.9 Nigella Saiiva 1OO.O Initia heienium 24.6 Ocimum Basilicum 2O2 Juniperus Communis 33.0 Ocimum Basilicum 2O2 Juniperus Communis 38.2 Ocimum Basilicum 32.8 Lactica Saiiva 44.5 Oenothera biennis linne 1OO.O Lactica Saiiva 50.7 Onobrychis viciafolia 45.0 Laportea Canadensis 30.2 Optainia sp. 33.4 Lathyrus Saivais 20.4 Origantin nationara 2O.S Lathyrus Saivais 52.5 Origantim vulgare 20.8 Lathyrus Sylvestris 27.7 Origantim vulgare 21.6 Lathyrus Sylvestris 36.8 Oryza sativa 42.4 Latiri is nobilis S2.O Oxyria digyna 57.0 Lavendula anguistifolia 26.4 Oxyria digyna 77.9 Lavendula anguistifolia 53.2 Panax quinquefoilis L. 23.5 Lavenditia iaiifolia 51.3 Panicum miliaceum 36.5 Ledum groenlandictim 44.4 Passiflora spp 35.8 Lentinus edodes 42.1 Passiflora spp 38.3 Lentinus edodes 1OO.O Passiflora spp 46.2 Lepidium sativum 44.2 Passiflora spp 1OO.O Levisticum officinale 20.8 Pastinaca Saiiva 21.7 Levisticum officinale 39.4 Pastinaca Saiiva 38.6 Lintin itsitatissini in 42.3 Pastinaca Saiiva 39.2 Litchi chinensis 25.7 Persea americana 32.5 Lolium multiflorum 20.6 Persea americana 38.6 Lolium perenne 28.7 Petasites Japonicus 26.2 Lonicera ranosissina 26.3 Phaiaris canariensis 80.0 Lonicera ranosissina 40.4 Phaseolus coccineus 44.4 Lonicera ranosissina 53.2 Phaseolus coccineus 79.1 Lonicera syringantha 95.8 Phaseolus mingo 27.0 Lotus corniculatus 1OO.O Phaseolus mingo 37.9 Lotus tetragonolubits 65.4 Phaseolus vulgaris 20.1 Linaria annia 55.7 Phaseolus vulgaris S1.9 Linaria annia 67.3 Phaseolus vulgaris 61.7 Lycopersicon esculenium 37.6 Phlox paniculata 22.9 Maius 31.8 Phlox paniculata 44.5 Maius 44.4 Phoenix dactylifera 29.6 Maius hupehensis (Pamp.) Rehd. 26.3 Physalis alkekengi 32.9 Maius hupehensis (Pamp.) Rehd. 67.0 Physalis ixocarpa 26.6 Maius sp. 65.3 Physalis ixocarpa 28.3 US 2007/01 22492 A1 May 31, 2007 51

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name (%) Latin name Stress Extract (%) Physalis pruinosa 27.3 Ribes Saiivum 48.2 Physalis pruinosa 47.8 Ribes Silvestre 26.3 Physalis pruinosa 93.1 Ribes Silvestre 1OO.O Physalis sp 39.1 Ribes tiva-crispa 57.5 Physalis sp 60.8 Rosa rugosa 27.8 Phytolacca americana 4.1.8 Rosa rugosa thunb. 37.5 Phytolacca americana 1OO.O Rosa rugosa thunb. 45.7 Phytolacca decandra syn. P. 85.9 Rosmarinum officinalis 44.2 americana Rosmarinum officinalis 65.9 Pimpinella anisum 2O2 Rubus Canadensis 45.5 Pimpinella anisum 68.4 Rubus idaeus 31.4 Pistin Saivitin 20.1 Rubus idaeus 57.2 Pistin Saivitin 25.8 Rubus idealis 28.5 Pistin Saivitin 27.0 Rubus idealis 38.0 Pistin Saivitin 51.8 Rubus occidentais 21.4 Plantago Coronopus 21.9 Rubus occidentais 36.5 Plantago Coronopus 48.6 Rubus occidentais 60.2 Plantago Coronopus 66.8 Rimes Scuttatus 84.5 Plantago major 35.1 Rumex crispiis line 52.5 Pleurotus spp 25.3 Rumex crispiis line 1OO.O Pleurotus spp 59.3 Rumex patientia 23.1 Pleurotus spp 85.2 Rumex patientia 65.8 Poa compressa 26.2 Rita graveolens 37.2 Poa praiensis 21.5 Sabal Serraiata syn. Serenoa repens 34.4 Poa praiensis 3O.O Sabal Serraiata syn. Serenoa repens 44.6 Podophyllum peltatum 33.9 Salix purpurea 67.8 Podophyllum peltatum SO.2 Salvia (elegens) S1.1 Polygonum aviculare linne 31.0 Sambucus Canadensis 44.8 Polygontin pennsylvanicum 56.6 Sambucus Canadensis 72.4 Polygontin persicaria 20.1 Sambucus Canadensis L. 67.8 Populus incrassata S4.9 Sambucus ebulus 44.3 Populus Tremula 31.0 Sanguisorba officinalis 1OO.O Populius X petrovskyana 1OO.O Santoina 37.9 Potentiiia anserina 22.1 Satureia montana 2O.O Potentiiia anserina 41.1 Satureia montana 21.3 Prints cerastis 30.1 Satureia repandra 36.3 Prunus persica 26.6 Scorzorera hipanica 27.1 Prunus persica 38.5 Scorzorera hipanica 31.7 Prunus spp 24.O Scuttellaria laterifiora 44.3 Prunus spp 49.1 Secale cereale 24.2 Psidium guaiaba 22.5 Secale cereale 31.1 Psidium guaiaba 44.3 Sechium edule 37.8 Psidium guaiaba 95.4 Sesamun indicum 59.2 Psidium spp 36.6 Setaria italica 33.0 Psidium spp 47.6 Silybum marianum 92.4 Psidium spp 87.6 Siim Sisartin 32.7 Pteridium aquilinum 22.O Siim Sisartin 33.1 Punica granathin S2.1 Siim Sisartin 813 Pyrus communis 39.5 Soianum melogena 21.9 Pyrus pyrifolia 33.7 Soianum melogena 26.1 Raphantis raphanistrain 24.5 Soianum melogena 34.O Raphantis raphanistrain 44.8 Soianum melogena 67.1 Raphantis raphanistrain 46.1 Soianum Tiiberosum 68.6 Raphantis Saivais 25.4 Solidago Canadensis 48.4 Raphantis Saivais 32.1 Solidago sp 31.4 Raphantis Saivais 38.1 Solidago virgau rea 56.2 Raphantis Saivais 63.6 Sorghum cafiorum 23.3 Raphantis Saivais 93.4 Sorghum dochna bicolor gir technicum 20.8 Reseda luteola 22.5 Sorghum dochna Snowdrew 21.4 Rhamnus fianglia 34.2 Sorghum dochna Snowdrew 27.7 Rhamnus fianglia 39.5 Spinacia oleracea 2S.O Rheum officinale 1OO.O Spinacia oleracea 32.1 Rheum palmatum 2O2 Spinacia oleracea 47.6 Rheum rhabarbarium 33.8 Spinacia oleracea 63.1 Rianits communis 20.9 Stachys affinis 31.7 Ribes nidigrolaria 44.5 Stachys affinis 1OO.O Ribes nidigrolaria 53.1 Stachys byzantina 30.9 Ribes nigrum 40.7 Stipa capiliata L. 20.1 Ribes nigrum L. SO.O Symphytum officinale 24.1 Ribes nigrum L. 60.1 Tanacetum cineranifolium 24.2 Ribes sativam Syme 47.9 Tanacetum cineranifolium 84.4 US 2007/01 22492 A1 May 31, 2007 52

TABLE 4-continued TABLE 4-continued Inhibition of MMP-9 by Plant Extracts Inhibition of MMP-9 by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Tanacetum vulgare R 25.7 Xanthium Sibiricum T S 38.6 Tanacetum vulgare S 75.6 Xanthium strumarium T S 33.5 Taraxacum officinale (Red ribe) S 21.1 Zea mayS T S 37.1 Teparty R 56.7 Zea mayS T O 65.5 Teucrium chamaedrys L. T R 27.3 Zingiber officinale T S 20.1 C. Floti,SStiltiS R 8. Zingiber officinale T W S8.9 Thymus herba-barona W 22.O Zingiber officinale T O 75.9 Thymus pseudoianuginostis R 36.8 Thymus pseudoianuginostis S 37.1 Thymus serpyllum S 26.O Thymus serpyllum W 42.7 0248) Thymus X citriiodorus O 22.7 Tiarella cordifolia R 1OO.O TABLE 5 Tragopogon porrifolius V 26.8 Tragopogon porrifolius O 28.4 Inhibition of HLE by Plant Extracts Tragopogon porrifolius S 42.1 Tragopogon sp. O 20.3 Inhibition Tragopogon sp. S 32.O Latin name Stress Extract (%) Tragopogon sp. W 66.3 Trichosanthes kirilowii O 66.5 Achillea millefolium A. O 21.9 Trifolium incarnatum R 47.9 Achillea millefolium A. S 24.5 Trifolium repens R 81.7 Aconium napeius A. O 25.8 Trigonelia foenim graecum S 39.6 Adiantum pedatum A. R 27.6 Triticale sp. O 64.1 Agrimonia eupatoria A. V 26.0 Trictim aestivitin W 24.5 Agropyron cristainin A. R 21.0 Trictim aestivitin S 29.4 Agropyron repens A. S 23.4 Triticum fungidumm S 35.8 Agropyron repens A. R 28.2 Triticum spelta S 34.7 Agropyron repens A. S 39.8 Tropaeoluim maints O 90.3 Agrostis Stofonifera A. O 38.9 Tropaeoluim maints W 64.4 Aichemia nois A. V 27.9 Tsiiga canOadensis O 21.5 Aichemia nois A. O 66.O Tsiiga canOadensis W 64.4 Aichemia nois A. R 1OOO Tsiiga diversifolia O 45.9 Aichemia nois A. S 23.5 Tsiiga diversifolia W 1OO.O Aikanna tinctoria A. S 26.2 Tsiiga F. macrophylia W 28.1 Aium Tiberosum A. S 57.9 Typha latifolia L. S 30.6 Aloe Vera A. O 2O.S Urtica dioica O 31.4 Ambrosia artemisiifolia A. O 29.1 Urtica dioica R 36.9 Amelanchier Sanguinea A. W 96.5 Urtica dioica S 41.7 Amelanchier Sanguinea A. V 52.4 Vaccinium anglisiifolium V 25.2 Anethlin graveolens A. O 32.1 Vaccinium anglisiifolium R 34.6 Anethlin graveolens A. W 22.8 Vaccinium anglisiifolium O 59.6 Angelica archangelica A. S 39.2 Vaccinium anglisiifolium R 65.7 Anthemis nobiis A. O 37.6 Vaccinium macrocarpon O 30.2 Anthemis nobiis A. S 26.4 Vaccinium macrocarpon S 39.0 Anthemistinctoria A. O 31.9 Vaccinium macrocarpon S 56.9 Anthemistinctoria A. S 38.4 Vaccinum macrocarpon V 39.2 Apium graveolens A. S 49.2 Vaccinum macrocarpon W 42.3 Arctii in minus A. O 46.4 Veratrum viride O 2O.S Arctostaphylos tiva-ursi A. R 1OOO Veratrum viride S 33.1 Aronia melanocarpa A. O 21.9 Verbascum thapsus S 43.1 Aronia melanocarpa A. W 78.4 Verbascum thapsus O 70.2 Aronia melanocarpa A. V 1OOO Veronica officinalis O 2O.S Aronia melanocarpa A. R 29.0 Viburnum trilobium Marsh. S 40.6 Aronia melanocarpa A. O 33.6 Vicia faba R 61.5 Artemisia dractinctius A. W 89.2 Vicia Saiva R 30.1 Ludoviciana A. O 33.4 Vigna angularia R 32.6 Ludoviciana A. S 20.7 Vigna angularia S 642 Aster sp A. R 26.2 Vignatinguiculata R 32.4 Beta vulgaris A. R 1OOO Vignatinguiculata O 47.4 Beta vulgaris spp. Maritima A. R 92.2 Vignatinguiculata S S1.O Borago officinalis A. S 22.6 Vinca minor S 21.3 Brassica naptis A. S 68.3 Vitis sp. V 28.3 Brassica naptis A. R 29.5 Vitis sp. O 29.4 Brassica nigra A. S 32.6 Vitis sp. S 45.4 Brassica oleracea A. O 22.9 Vitis sp. V 50.7 Brassica oleracea A. V 20.8 Vitis sp. W 61.6 Brassica oleracea A. R 22.2 Vitis sp. R 1OO.O Brassica rapa A. S 23.2 Weigela coracensis W 35.5 Brassica rapa A. R 26.9 Withania somnifera S 35.5 Bromits inermis A. O 34.1 US 2007/01 22492 A1 May 31, 2007 53

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Bromits inermis 21.9 Lavandhiia iaiifolia 42.2 Calamintha nepeta 35.4 Leonirits cardiaca 1OOO Canna editiis 56.4 Lepidium sativum 1OOO Canna editiis 21.4 Lolium multiflorum 31.0 Cartin carvi 24.2 Lolium perenne 20.8 Chaerophyllum bulbosum 25.5 Lolium perenne 21.7 Chenopodium bonus-henrict is 24.0 Lolium perenne 22.1 Chenopodium bonus-henricus 85.8 Malva Sylvestris 22.9 Chenopodium quinoa SO4 Matricaria recuita 28.5 Chrysanthemum coronarium 26.0 Melaleuca alternifolia 21.9 Cicer arrieintin 23.3 Melissa officinalis 23.4 Cichorium intybus 32.1 Mentha piperita 31.6 Citritius ianatus 26.3 Mentha piperita 33.2 Coix Lacryma-Jobi 66.1 Mentha pullegitim 42.2 Cosmos Sulphureus 38.8 Mentha pullegitim 21.5 Cosmos Sulphureus 20.7 Mentha pullegitim 33.8 Crataegus sp 84.1 Mentha spicaia 24.3 Crataegus sp 23.6 Oenothera biennis 25.2 Crataegus sp 21.7 Oenothera biennis 78.8 Crataegus submolis 34.0 Origantin majorana 37.4 Cryptotaenia Canadensis 22.1 Oxyria digyna 28.2 Cuctimis anguria 26.2 Panicum miliaceum 33.3 Cuctimis Angliria 53.4 Peucedanum cervaria 23.4 Cucumis meio S3.6 Phaiaris arundinacea 22.4 Ciclinis Saiivus 53.3 Phaiaris canariensis 27.8 Curcuna Zedoaria 24.3 Phaseolus coccineus 28.3 Cymbopogon citratus 91.2 Phaseolus mingo 37.8 Datisca Cannabina 55.7 Phaseolus vulgaris 24.3 Daiiciis Caroia 1OOO Phaseolus Vulgaris 74.3 Daiiciis Caroia 24.7 Phleum pratense 27.8 Daiiciis Caroia 37.9 Physalis ixocarpa 21.5 Digitalis purpurea 34.0 Physalis Ixocarpa 26.5 Dirca palustris 20.3 Physalis Pruinosa 6O2 Dirca palustris 27.9 Phytolacca americana 1OOO Doichos Labiah 21.5 Plantago Coronopus 21.1 Dryopteris filix-mas 58.8 Plantago Coronopus 25.7 Dryopteris filix-mas 22.0 Plantago major 26.0 Echinacea purpurea 38.2 Plectranthus sp. 23.1 Echinacea purpurea 28.1 Poa pratensis 21.7 Eleusine Coracana 20.7 Polygonum aviculare 79.7 Erigeron Canadensis 29.6 Portuiaca oievcae 34.5 Fagopyrim esculenium 29.3 Poterium sanguisorba 25.8 Fagopyri in tatarictim 24.4 Poterium sanguisorba 34.6 Foeniculum vulgare 25.1 Poterium sanguisorba 31.0 Fragaria Xananassa 22.3 Pteridium aquilinum 544 Fragaria Xananassa 1OOO Raphantis Saivais 66.4 Fragaria Xananassa 21.4 Raphantis Saivais 81.8 Fragaria Xananassa 29.4 Rheum officinale 37.9 Fragaria Xananassa 21.6 Ribes nigrum 1OOO Gainsoga ciliata 61.6 Ribes nigrum 47.6 Gaium odoratum 21.0 Ribes nigrum 27.5 Gaultheria hispidula 33.7 Ribes rubrum 35.4 Gentiana littea 52.1 Ribes Sylvestre 1OOO Giechona hederacea 21.8 Rosa rugosa 95.1 Glycine Max 813 Rosa rugosa 24.6 Glycyrrhiza glabra 1OOO Rosmarinus officinalis 58.4 Glycyrrhiza glabra 6.3.3 Rubus idaeus 27.6 Guizotia abyssinica 36.9 Rubus idaeus 33.0 Hamamelis virginiana 1OOO Rubus idaeus 27.9 Heianthus Tiiberosus 32.1 Rubus idaeus 37.4 Heliotropium arborescens 22.8 Rumex Acetosa 45.2 Heliotropium arborescens 24.9 Rumex crispiis 26.1 Helleborus niger 25.6 Rumex crispiis 1OOO Hordeum vulgare S8.1 Rumex Scittatus 43.8 Hyperictim perforatin 24.8 Rita graveolens 28.7 Hyssopus officinalis 21.1 Saccharum officinarum 29.6 Hyssopus officinalis 93.6 Saccharum officinarum 23.8 Lactuca serioia 34.3 Salvia elegans 1OOO Latiri is nobilis 1OOO Salvia officinalis 95.7 Lavandhiia iaiifolia 57.1 Salvia officinalis 77.9 Lavandhiia iaiifolia 43.7 Salvia officinalis 83.7 US 2007/01 22492 A1 May 31, 2007 54

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Salvia officinalis 2O.S Arctostaphylos tiva-ursi 33.1 Saivia Sciarea 1OOO Arctostaphylos tiva-ursi 1OOO Saivia Sciarea 28.6 Arctostaphylos tiva-ursi 23.4 Santolina Chamaecyparissus 27.1 Armoracia rusticana 22.5 Satureia montana 23.2 Aronia melanocarpa 79.0 Satureia montana 27.7 Aronia melanocarpa 1OOO Scorzonera hispanica 60.1 Aronia melanocarpa 22.7 Scutellaria laterifiora 45.9 Aronia melanocarpa 29.6 Senecio vulgaris 34.0 Artemisia absinthium 31.5 Sonchi is oleracetis 29.1 Artemisia absinthium 24.2 Sorghum dochna 21.1 Aster 29.2 Sorghum dochna 24.4 Beckmannia eruciformis 22.7 Sorghum durra 23.4 Beta vulgaris 1OOO Sorghum durra 23.6 Betula glandulosa 26.7 Spinacia oleracea 26.8 Borago officinalis 25.7 Stellaria graminea 24.8 Brassica Naptis SO4 Symphytum officinale 91.6 Brassica naptis 48.2 Tanacetum cinerariifolium 28.3 Brassica nigra 23.9 Tanacetum vulgare 46.3 Brassica oleracea 28.1 Tanacetum vulgare 33.7 Brassica oleracea 22.5 Taraxacum officinale 26.4 Brassica rapa 56.4 Taraxacum officinale 24.0 Calamintha nepeta 24.8 Taraxacum officinale 21.0 Calamintha nepeta 38.8 Teucrium chamaedrys 37.0 Canna eduis 66.3 Thymus fragantissimils 2O2 Capsella bursa-pastoris 25.8 Thymus herba-barona 20.8 Carthamus tinctorius 22.2 Thymus vulgaris 77.9 Chelidonium maius 31.6 Thymus vulgaris 23.6 Chenopodium album 21.3 Thymus x citriodorus 21.3 Cichorium endivia subsp. Endivia 21.4 Thymus x citriodorus 21.1 Cicer arrieintin 50.7 Trichosanthes kirilowii 23.2 Cichorium endivia subsp. Endivia 48.5 Trigonelia foenim graecum 32.O Cichorium endivia subsp. Endivia 27.9 Triticum durum 22.0 Coix Lacryma-Jobi 24.5 Triticum turgidium 6O.O Cornus Canadensis 36.1 Triticum spelta 47.6 Crataegus sp 57.8 Urtica dioica 33.3 Cucurbita Pepo 23.1 Vaccinium augustifolium 42.6 Curcuna Zedoaria 24.0 Vaccinium Corymbosum 22.4 Datura metel 21.0 Vaccinium Corymbosum 21.6 Daiiciis Caroia 32.3 Vaccinium macrocarpon 22.5 Daiiciis Carroia 90.9 Vaccinium macrocarpon 54.8 Dipsactis sativus 32.7 Valerianeia iocusia 49.2 Dirca palustris 33.5 Veronica officinalis 43.7 Doichos Labiah 32.1 Viburnum trilobium Marsh. 75.4 Dryopteris filix-mas 80.9 iiis 33.8 Echinacea purpurea 63.0 iiis 1OOO Elymus incetts 25.9 iiis 21.0 Erigeron Canadensis 43.O Zea Mays 95.2 Erigeron speciosus 22.8 Achillea millefolium 28.8 Erigeron speciosus 24.2 Achillea millefolium 27.3 Erysimum perofskianum 20.8 Aconium napeius 23.1 Fagopyrim esculenium 32.9 Aconium napeius 97.7 Fagopyri in tatarictim 41.2 Acorus calamus 2O.O Focniculum vulgare 25.7 Adiantum pedatum 1OOO Foeniculum vulgare 42.5 Agastache foeniculum 25.3 Foeniculum Vulgare 24.1 Ageratum Conyzoides 28.5 Gainsoga ciliata 2SO Agropyron cristainin 37.3 Gaium odoratum 89.4 Agropyron repens 31.4 Gaultheria hispidula 35.1 Aichemilia nois 20.6 Gaultheria hispidula 67.2 Aichemilia nois 56.1 Gaultheria procumbens 74.7 Aichemilia nois 28.1 Glycine max 24.6 Aichemilia nois 25.3 Glycyrrhiza glabra 56.8 Allium cepa 2O2 Glycyrrhiza glabra 3O.O Aium sativum 1OOO Glycyrrhiza glabra 92.4 Aium tuberosum 1OOO Glycyrrhiza glabra 28.6 Althaea officinalis 30.8 Hamamelis virginiana 1OOO Amaranthus caudatus 22.3 Hamamelis virginiana 29.3 Amelanchier Sanguinea 88.3 Hedeona pullegioides 6O.O Anethlin graveolens 26.2 Helenium hoopesii 37.3 Angelica archangelica 43.2 Helenium hoopesii 34.7 Anthemis nobiis 21.7 Heianthus tuberosus 21.4 US 2007/01 22492 A1 May 31, 2007 55

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Helichrysum thianschanicum 43.O Polygonum aviculare 25.4 Helichrysum thianschanicum 39.2 Polygontin pensylvanictim 21.3 Heliotropium arborescens 22.8 Portuiaca oleracea 28.0 Heliotropium arborescens 39.5 Poterium sanguisorba 25.6 Helleborus niger 34.2 Poterium sanguisorba 21.9 Hypericum henryi 23.7 Prunella vulgaris 23.4 Hyperictim perforatin 23.8 Pteridium aquilinum 43.1 Hyssopus officinalis 45.1 Reseda Odorata 46.5 Hyssopus officinalis 24.2 Rhaphant is sativus 32.6 Initia heienium 96.2 Rheum x cuitorum 20.9 Ipomoia batatas 21.9 Ribes nidigrolaria 29.8 Lactica Saiiva 35.1 Ribes nidigrolaria 53.7 Laportea Canadensis 25.1 Ribes nigrum 20.3 Laportea Canadensis 26.5 Ribes Silvestre 91.6 Laserpitium laiifolium 22.1 Ricinits communis 46.0 Lathyrus Saivais 29.9 Rosmarinus officinalis 60.4 Lathyrus Saivais 27.8 Rubus idaeus 28.2 Lathyrus Saivais 28.1 Rubus occidentais 93.6 Latiri is nobilis 1OOO Rubus occidentais 40.O Lavandhiia angustifolia 65.7 Rumex acetoseiia 24.3 Ledum groenlandictim 1OOO Rumex crispiis 1OOO Leonori is cardiaca 61.3 Rumex patientia 32.O Lepidium sativum 1OOO Rumex Scittatus 28.6 Levisticum officinale 91.4 Rita graveolens 23.4 Lolium perenne 37.3 Saccharum officinarum 30.2 Lotus tetragonolobits 21.8 Salix purpurea 24.8 Lupinus polyphyllius 423 Salvia elegans 1OOO Maius hupehensis 25.9 Salvia officinalis 52.4 Medicago Saiiva 32.1 Salvia officinalis 1OOO Melaleuca alternifolia 40.O Salvia officinalis 1OOO Melissa officinalis 23.1 Saivia Sciarea 1OOO Mentha arvensis 65.5 Saivia Sciarea 23.0 Mentha piperita 24.2 Saivia Sciarea 31.1 Mentha piperita 23.7 Sambucus ebulus 52.1 Mentha piperita 34.2 Sambucus ebulus 48.6 Mentha pullegitim 6.3.3 Sanguisorba officinalis 1OOO Mentha pullegitim 30.2 Santolina Chamaecyparissus 1OOO Mentha spicaia 45.9 Serrainia tinctoria 56.8 Monarda didyma 47.7 Satureia montana 34.1 Nepeta cataria 1OOO Scolymits hispanicus 37.9 Nicotiana tabacum 75.8 Scutellaria laterifiora 54.7 Hordeum vulgare subsp. Vulgare 33.4 Senecio vulgaris 35.3 Ocimum basicum 40.1 Solidago sp 22.6 Ocimum basicum 27.9 Sonchi is oleracetis 23.7 Oenothera biennis 26.3 Sorghum cafiorum 27.1 Oenothera biennis 1OOO Sorghum dochna 40.7 Oenothera biennis 49.6 Sorghum dochna 21.4 Oenothera biennis S4.O Sorghum Sudanense 23.3 Origantim vulgare 1OOO Sorghum Sudanense 92.9 Origantim vulgare 26.7 Stellaria graminea 25.4 Origantim vulgare 21.3 Steiaria media 3O4 Oryza Sativa 34.5 Steiaria media 22.0 Oxalis Deppei Lodd. 27.4 Tanacetum vulgare 57.3 Panicum miliaceum 25.3 Tanacetum vulgare 38.4 Pastinaca Saiiva 95.0 Tanacetum vulgare 38.2 Petroselinum crispum 445 Tanacetum vulgare 26.3 Petroselinum crispum 26.5 Taraxacum officinale 2O.O Peucedanum cervaria 25.1 taraxacum officinale 28.0 Phaseolus coccineus 30.9 Thymus fragantissimils 79.9 Phaseolus coccineus 27.5 Thymus fragantissimils 26.2 Phaseolus mingo 24.3 Thymus herba-barona 2O2 Phlox paniculata 37.9 Thymus serpyllum 22.2 Physalis pruinosa 26.5 Triticosecale spp. 29.7 Phytolacca americana 1OOO Triticum durum 37.8 Pimpinella anisum 23.7 Triticum spelta 31.0 Plantago Coronopus 25.1 Triticum spelta 37.9 Plantago major 2SO Typha latifolia 27.5 Plantago major 2O.S Urtica dioica 60.3 Plantago major 23.6 Vaccinium corymbosum 33.2 Poa compressa 28.5 Vaccinium anglisiifolium 43.7 Poa praiensis 37.5 Vaccinium macrocarpon 57.8 US 2007/01 22492 A1 May 31, 2007 56

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Vaccinium macrocarpon 59.9 Capsicum annittin 22.6 Valerianeia iocusia 32.1 Carex morrowii 26.0 Veratrum viride 22.1 Carex morrowii 49.8 Verbascum thapsus 33.8 Carya cordiformis 28.8 Viburnum trilobium 21.3 Carya cordiformis 21.0 Viburnum trilobium 73.0 Carya cordiformis 88.7 Vicia faba 21.2 Ciennais armandi 20.1 Vignatinguiculata 20.1 Chaerophyllum bulbosum 22.8 iiis 26.0 Chaerophyllum bulbosum 24.3 iiis 66.1 Agarict is bisporatiis 25.4 iiis 41.7 Chelidonium maius 39.0 iiis 30.7 Chenopodium bonus-henricus 44.3 Xanthium Sibiricum 22.1 chrysanthemum coronarium 33.4 Zea mayS 20.3 chrysanthemum coronarium 23.9 Abies lasiocarpa 22.4 Cichorium endivia subs. Endivia 44.3 Achillea millefolium 21.1 Cichorium endivia subs. Endivia 2O.S Aconium napeius 1OOO Circium arvense 49.7 Acorus calamus 21.0 Citrulius colocynthis 37.0 Ageratum Conyzoides 20.1 Citrulius colocynthis 35.5 Agrimonia eupatoria 59.6 Citrus innettoides 47.1 Agropyron cristainin 53.4 Citrus innon 26.2 Agropyron repens 22.6 Citrus innon 73.9 Agrostis alba 25.3 Citrus Sinensis 25.2 Aichemilia nois 88.7 Coix Lacryma-Jobi 32.7 Aichemilia nois 42.6 Coix Lacryma-Jobi 31.4 Aichemilia nois 704 Corchorus oitorius 24.4 Aichemilia nois 31.2 Cornus Canadensis 41.3 Allium ascalonictim 42.9 Crataegus sp 34.0 Aium sativum 1OOO Crataegus submolis 39.6 Aium tuberosum 1OOO Circlina longa 55.3 Alpinia officinarim 21.9 Curcuna Zedoaria 24.4 Alpinia officinarim 1OOO Cydonia oblonga 35.2 Amaranthus candatus 36.0 Cynara Scolymits 41.2 Amaranthus gangeticus 66.8 Cynara Scolymits 36.8 Ananas Conostis 20.3 Dactiis Giomerata 31.9 Ananas Conostis 23.8 Daitira Stranonium 25.9 Anethlin graveolens 35.8 Daiiciis Caroia 92.3 angelica archangelica 53.5 Daiiciis Caroia 31.0 Anthemis nobiis 45.3 Dipsactis sativus 1OOO Anthemistinctorium 47.5 Dirca palustris 31.4 Anthriscus cerefolium 2O.S Doichos iabiab 23.1 Arctii in minus 54.1 Dryopteris filix-mas 68.2 Arctostaphylos tiva-ursi 28.1 Echinacea purpurea 38.2 Arctostaphylos tiva-ursi 1OOO Eleusine Coracana 22.1 Aronia melanocarpa 1OOO Elymus incetts 37.9 Aronia melanocarpa 42.7 Erigeron speciosus 3S.O Aronia prunifolia 39.0 Erysimum perofskianum 22.6 Artemisia absinthium 25.6 Erysimum perofskianum 23.2 Artemisia dractinius 31.3 Fagopyrim esculenium 24.7 Artemisia dractinius 22.3 Foeniculum vulgare 31.4 Aster 20.9 Foeniculum vulgare 69.1 Avena Saiiva 1OOO Foeniculum vulgare 38.5 Averrhoa caramboia 25.8 Fragaria x ananassa SO4 Beta vulgaris 1OOO Fragaria x ananassa 30.2 Beta vulgaris 59.3 Fragaria x ananassa 28.4 Beta vulgaris 41.4 Passifiora spp. 30.2 Betula glanditiosa 618 Passifiora spp. 59.4 Boesenbergia rotunda 36.9 Passifiora spp. 24.4 Boesenbergia rotunda 42.5 Fictis vesiculosis 42.7 Boletus eduis 43.1 Gainsoga ciliata 49.3 Borago officinalis 36.3 Gaultheria hispidula 36.9 Brassica hiria 30.2 Gentiana macrophylla 26.1 Brassica iuncea 41.4 Ginkgo biloba 27.1 Brassica Naptis 29.9 Glycyrrhiza glabra S8.1 Brassica naptis 22.9 Glycyrrhiza glabra SO4 Brassica oleracea 25.6 Glycyrrhiza glabra 25.1 Brassica oleracea 27.0 Gossypium herbaceum 22.7 Brassica oleracea 26.5 Gossypium herbaceum 27.3 Brassica rapa 24.8 Guizotia abyssinica 38.5 Bromits inermis 27.8 Hamamelis virginiana 37.1 Canna editiis 40.3 Hamamelis virginiana 1OOO US 2007/01 22492 A1 May 31, 2007 57

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Hedeona pullegioides 28.5 Oenothera biennis 24.2 Hedeona pullegioides 28.2 Oenothera biennis 58.6 Helenium hoopesii 31.7 Onobrychis viciifolia 42.6 Helenium hoopesii 56.0 Origantim vulgare 53.8 Heianthus tuberosus 23.7 Oryza sativa 33.3 Helichrysum thianschanicum 38.4 Oxalis Deppei 30.8 Helichrysum thianschanicum 27.0 Panicum miliaceum 21.2 Helleborus niger 32.1 Pastinaca Saiiva 53.9 Schizonepeta tenuifolia 29.1 Pastinaca Saiiva 20.8 Schizonepeta tenuifolia 21.1 Pastinaca Saiiva 26.9 Hibiscus cannabinus 39.9 Petroselinum crispum 58.2 Hibiscus cannabinus 21.1 Phaseolus coccineus 27.1 Huntilus lupuits 54.8 Phaseolus vulgaris 37.9 Huntilus lupuits 50.5 Phaseolus vulgaris 22.2 Hydrastis Canadensis 20.9 Phaseolus vulgaris 23.2 Hypericum henryi 32.5 Phlox paniculata 21.3 Hyperictim perforatin 27.9 Physalis pruinosa 35.2 Hyperictim sp 55.9 Phytolacca americana 1OOO Hypomyces lactifiutorum 42.7 Plantago Coronopus 21.2 Iberis amara 1OOO Plantago Coronopus 48.2 Initia heienium 30.1 Poa pratensis 50.7 Ipomoia batatas 27.4 Podophyllum peltatum 27.9 Ipomoia batatas 44.9 Polygonum chinense 2SO Juniperus Communis 57.8 Polygonum aviculare 26.0 Laportea Canadensis 63.5 Polygonum aviculare 1OOO Latiri is nobilis 73.6 Polygontin pensylvanictim 423 Latiri is nobilis 21.2 Polygontin persicaria 28.8 Lavandula angustifolia 22.7 Populus incrassata 1OOO Lavandhiia angustifolia 25.1 Populus Tremula 48.5 Lavandhiia iaiifolia 1OOO Populus x petrowskyana 44.1 Lavandhiia iaiifolia 28.5 Populus x petrowskyana 1OOO Ledum groenlandictim S4.3 Populus x petrowskyana 72.0 Lentinus edodes 25.7 Portuiaca oleracera 33.7 Leonirits cardiaca 24.3 Poterium sanguisorba 1OOO Lepidium sativum 1OOO Prints spp. 39.6 Levisticum officinale 41.2 Prunus persica 21.4 Litchi chinensis 1OOO Prunus persica 26.6 Lolium multiflorum 24.0 Psidium guaiava 37.7 Lolium perenne 27.8 Psoralea corylifolia 51.5 Lonicera ranosissina 20.9 Pteridium aquilinum 76.2 Lupinus polyphyllius 35.1 Pteridium aquilinum 27.9 Lupinus polyphyllius 2O.S Punica granathin 66.4 Luzula Sylvatica 22.6 Rehmannia glittinosa 83.0 Majorana hortensis 20.1 Frangula aint is 40.7 Maius spp. 37.8 Raphantis Saivais 36.5 Maius spp. 45.1 Raphantis Saivais 22.4 Maius hupehensis 24.4 Reseda luteola 23.6 Melaleuca alternifolia 26.7 Reseda Odorata 20.3 Melissa officinalis 20.7 Frangula aint is 65.3 meniha arvensis 34.0 Rheum officinale 1OOO Mentha piperita 60.1 Rheum officinale 33.3 Mentha pullegitim 24.5 Rheum x cuitorum 34.0 Mentha pullegitim 24.8 Ricinits communis 27.5 Mentha spicaia 24.4 Ribes Grossuiaria 24.8 Mentha suaveolens 28.9 Ribes nidigrolaria 24.4 Monarda didyma 54.7 Ribes nigrum SO.1 Musa paradisiaca 21.4 Ribes nigrum 23.8 Musa paradisiaca 32.8 Ribes nigrum 64.1 nastairtium officinale 1OOO Ribes Sylvestre 32.4 Nepeta cataria 60.1 Rosa rugosa 1OOO Nepeta cataria 23.4 Rosmarinus officinalis 75.8 Nigella Saiiva 23.2 Rosmarinus officinalis 46.6 Agarict is bisporatiis 25.8 Rubus idaeus 27.6 Psidium spp. 28.3 Rubus idaeus 24.3 Pleurotus spp. 31.6 Rubus idaeus 35.5 Citrus reticulata 32.7 Rubus occidentais 93.2 Citrus reticulata 29.4 Rubus occidentais 42.1 Ocimum Basilicum 30.7 Rubus occidentais 2O.S Ocimum Basilicum 30.9 Rumex acetoseiia 44.9 Ocimum Basilicum 39.1 Rumex crispiis 31.3 Oenothera biennis 29.6 Rumex crispiis 1OOO

US 2007/01 22492 A1 May 31, 2007 59

TABLE 5-continued TABLE 5-continued Inhibition of HLE by Plant Extracts Inhibition of HLE by Plant Extracts Inhibition Inhibition Latin name Stress Extract (%) Latin name Stress Extract (%) Oenotherafiniticosa spp. 109.8 Begonia mannii 82.5 Oenotherafiniticosa spp. 61.3 Begonia mannii 72.8 Oenotherafiniticosa spp. 97.5 Begonia polygonoides 79.0 Oenotherafiniticosa spp. 105.9 Begonia polygonoides 74.8 Veronica austriaca Ssp teucrium 68.6 Begonia polygonoides 73.2 Veronica austriaca Ssp teucrium S8.1 Fushia spp. 76.6 Coreopsis verticiliata 55.6 Fushia spp. 70.7 Coreopsis verticiliata 704 Fushia spp. 76.9 Potentilla fruticosa 104.8 Button is timbellatus 58.8 Potentilla fruticosa 99.4 Onociea sensibiis 54.7 Potentilla fruticosa 98.6 Onociea sensibiis SO.1 Vernonia gigantea SO4 Pinus cembra 83.2 Vernonia gigantea 62.3 Pinus cembra 76.3 Vernonia gigantea 51.2 Corntis sericea 104.O Vernonia gigantea 50.7 Corntis sericea 53.4 Penstemon digitalis 64.5 Corntis sericea 91.8 Penstemon digitalis 63.5 Corntis sericea 51.0 Penstemon digitalis 57.3 Corntis sericea 98.5 Penstemon digitalis 63.4 Hydrangea quiercifolia S8.1 Penstemon digitalis 67.8 Solidago Caesia 60.7 Maius spp. 56.1 Solidago Caesia 6O.S Maius spp. 56.7 Cornus aiba 98.9 Maius spp. SO.8 Cornus aiba 106.7 Maius spp. 51.2 Cornus aiba 85.3 Hosta Sieboidiana SO.9 Carpinnis Caroliniana 95.4 Hamameis mois 99.1 Carpinnis Caroliniana 86.2 Hamameis mois 94.1 Carpinnis Caroliniana 94.5 Hamameis mois 89.4 Astilbe chinensis S4.3 Chaenomeles x superba 56.2 Astilbe chinensis S.O.3 Chaenomeles x superba 71.9 SymphoricarpoS albits 52.O Chaenomeles x superba 66.6 Euphorbia amygdaloides 103.8 Chaenomeles x superba 52.O Euphorbia amygdaloides 75.2 Centatinea dealbata SO.9 Euphorbia amygdaloides 71.3 Centatinea dealbata 74.1 Viburnum plicatum 61.O Paeonia spp. A. 79.8 Viburnum plicatum 57.9 Paeonia spp. 58.6 Buxus microphylla 58.0 Paeonia spp. 79.6 Astillboides tabularis 104.2 Paeonia spp. 58.5 Astillboides tabularis 108.1 Paeonia spp. 82.O Astillboides tabularis 100.3 Paeonia spp. 6O.O Staphylea trifolia 63.6 Lysimachia clethroides 83.3 Bergenia x Schmidtii 10O.S Lysimachia clethroides 64.3 Bergenia x Schmidtii 113.7 Lysimachia clethroides 85.8 Bergenia x Schmidtii 99.3 Lysimachia clethroides 67.8 Rodgersia podophylia 68.9 Magnolia x loebneri 61.4 Rodgersia podophylia 59.4 Iberis sempervirens 624 Rodgersia podophylia 56.5 Iberis sempervirens 63.8 Geranium phaeum 92.7 Filipendula vulgaris 98.3 Geranium phaeum 84.3 Filipendula vulgaris 94.5 Geranium phaeum 101.0 Filipendula vulgaris 96.3 Rubits pubescens 71.5 Geranium sanguinet in 89.4 Rubits pubescens 76.2 Geranium sanguinet in 6.3.3 Rubits pubescens 82.8 Geranium sanguinet in 82.6 Taxis x media 60.1 Geranium sanguinet in 53.2 Taxis x media 61.6 Garanium Sanginet in 88.8 Taxis x media 52.3 Garanium Sanginet in 57.7 Geranium X cantabrigiense 106.1 Philadelphus coronarius 55.5 Geranium X cantabrigiense 94.2 paeonia Stiffiniticosa S8.9 Geranium X cantabrigiense 95.9 paeonia Stiffiniticosa 52.1 Fuchia magellanica e 10O2 Paeonia suffiniticosa 73.8 Fuchia magellanica 91.9 Paeonia suffiniticosa 52.2 Fuchia magellanica 102.2 Paeonia suffiniticosa 58.7 Microbiata decussaia 51.5 Paeonia suffiniticosa SO4 Microbiata decussaia 51.9 Dahlia spp. 77.4 Rhododendron spp. 51.2 Begonia convolvulacea 69.8 Stephanandra incisa 102.5 Begonia convolvulacea 67.5 Stephanandra incisa 104.6 Begonia convolvulacea 72.6 Stephanandra incisa 99.1 Begonia emini 72.8 Corylus maxima SO.8 Begonia emini 77.2 Corylus maxima 57.1 Begonia emini 75.4 Cypertis alternifolius 56.2 Begonia glabra 82.3 Soleiroia Soleiroi 51.2 US 2007/01 22492 A1 May 31, 2007 60

0251 1. Analytical Scale Extraction Selection of TABLE 5-continued Plants/Extracts Inhibition of HLE by Plant Extracts 0252) The processed plant materials (leaves, roots, seeds and the like) were obtained by dedicated greenhouse culti Inhibition Latin name Stress Extract (%) vation (with or without physical/chemical stress), from commercial Suppliers, or by gathering from non-cultivated Soleiroia Soleiroi G R 68.0 Streitzia reginae T R 106.5 natural sources. For each plant used in either analytical scale Streitzia reginae A. R 94.3 or large Scale extraction, a properly identified and labelled Streitzia reginae G R 111.7 sample was kept in storage in the laboratory. Hedychium coronarium T R 53.5 Hedychium coronarium A. R 86.9 0253) The extraction protocols for both the preliminary Hedychium coronarium G R 74.6 Streitzia reginae T R 78.6 analytical scale and large Scale extractions are shown gen Streitzia reginae A. R 78.0 erally in FIG. 6. Streitzia reginae G R 107.3 SymphoricarpoS orbiculatus G R 58.7 0254 The collected dried plant material (2-10 g) was first Rodgersia spp. A. R 59.5 Submitted to Solid-liquid extractions to generate crude Rodgersia spp. G R 59.0 Lamiastrum galeobdolon R 91.5 extract A (mg scale). Two different solvents were tested Astilbe x arendsii A. R 84.5 (ethanol/methanol or ethanol/water mixtures). The extracts Cienais alpina A. R 544 were then defatted with hexane to yield hydrbalcoholic or Stewartia pseudocamelia R 75.5 Stewartia pseudocamelia A. R 84.1 alcoholic extract B and hexane extract C. A partitioning of Stewartia pseudocamelia G R 813 extract B with ethyl acetate was then performed after dilu Pinus mugo R S8.9 tion with water to yield aqueous extract E and organic Pinus mugo A. R 53.7 Pinus mugo G R 61.7 extract F. Rubus thibetanus R 97.6 Rubus thibetanus A. R 97.9 0255 The extracts were sampled and evaluated for their Rubus thibetanus G R 95.4 ability to inhibit one or more target protease and for their Rubus arcticus R 89.3 ability to affect one or more cellular activity in the skin using Rubus arcticus A. R 85.5 Rubus Phoenicolasius G R 93.2 the methods described below. ribes americanum R 704 Passifiora spp. O 624 0256 Analysis of the results allows for the selection of Rubus occidentais R 70.9 plant materials for the large-scale extraction. The selection Nicotiana tabacum G O 60.9 includes a decision regarding part of the plant and quantity Beta vulgaris O 71.3 of dried material needed to obtain sufficient mass of extract for pure active compound isolation. The selection also involves a choice of solvent system (aqueous versus alco Example III holic) and active extract (B, E or F) to be used in further work. Exemplary Purification of Inhibitory Activity Found 0257 The extracts were also analyzed by Thin Layer in an Extract Chromatography (TLC) with different reagents specific to 0249 Extracts were separated by HPLC on an Agilent classical chemical groups of natural products (terpenes, 1100 system (San Fernando, Calif.). Briefly, 100 uL of a alkaloids, phenolic acids, polyphenols) to evaluate the increase in concentration achieved by partitioning at each crude extract prepared as described in Example I was step, and also to remove any materials likely to produce false applied on a C18 reverse-phase column (Purospher RP-185 positive results (fatty acids, chlorophylls) and to provide an um, 4.0x125 mm (HP), Agilent, San Fernando, Calif.). indication of which fractionation steps to use in further Elution of compounds was achieved with a linear gradient of extractions. 10-85% acetonitrile. Fractions were collected, evaporated, resuspended in aqueous buffer and then reanalysed for their 0258 2. Large Scale Extraction Isolation inhibition activity on specific enzymes as already described. 0259 For each new specimen, a repeat analytical scale extraction is performed to confirm the biological activity Fractions of interest (demonstrating a biological activity) before beginning the large-scale extraction process. were then re-isolated at a larger scale for further analysis and characterisation. 0260 The first step is to release the secondary metabo lites from the dried and powdered material by means of an Example IV all purpose solvent mixture which is selected based on the results obtained in the analytical scale preparation. This can Preparation of Plant Extracts (Method B) be done by Successive maceration/percolation operations using the same solvent which should dissolve most natural 0250 Method B is summarized in general terms in FIG. compounds at the same time. The bulk of the inert and 5. The method can be divided into two main parts corre insoluble material such as cellulose is then removed by sponding to preliminary analytical scale extraction and a filtration. Conditions of drying and grinding are controlled second larger scale extraction process. (temperature of drying less than 45° C., particles size). US 2007/01 22492 A1 May 31, 2007 61

0261) The second step is to remove a portion of the 0266 All the extracts are analyzed by TLC to compare unwanted material in a series of liquid-liquid low resolution with analytic scale extracts. extractions using solvents of different polarity with the aim of a multi-gram mixture containing all the natural products Example V of interest and to remove the most of the undesired material. 0262 The extraction protocol is illustrated in FIG. 6 and Protease Inhibition by Plant Extracts in a Human is essentially the same as the procedure for the analytical Skin Model preparation. The dried and pulverized material (2-3 Kg for 0267 A cellular model of the skin was used to determine large scale) is extracted repeatedly (maceration/percolation) the potential inhibitory effect of aqueous and ethanolic plant with ethanol/methanol 85:15 V/v (a) or ethanol/water extracts prepared as described in Example I in the skin. 85:15 V/v (b) mixtures (3x5-10 L) at room temperature for Human dermal fibroblasts (Cascade Biologics, 5x10/well), 2x24–48 h, based on the analytical scale results (yield of type 1 collagen (3 mg/ml. Sigma), and cell culture medium extraction). were pipetted into 12 or 24-well untreated Falcon plates and incubated for 1 hour at 37° C., allowing for gel formation. 0263. In the case of an alcoholic extraction (a), the Cell culture medium was then added to the wells and the gels combined alcoholic extracts (A) are concentrated under were incubated overnight at 37°C. in a 5% CO controlled reduced pressure, diluted with water (10-15%) and extracted atmosphere. The gels were incubated for 5 days, with media with hexane (or heptane) to yield hexane extract (C) and changes at days 2 and 4, allowing for fibroblast proliferation, hydroalcoholic fraction (B). This is then concentrated and with collagen and protease synthesis and secretion into the diluted with ethanol (20%) before being extracted with ethyl gel. On day 5, the media were removed and donor-matched acetate to yield aqueous (E) and ethyl acetate extracts (F). human epidermal keratinocytes (Cascade Biologics, 10 cells/well) in biological medium were gently pipetted onto 0264. In the case of an hydroalcoholic extraction (b), the the gels. The wells were further incubated for 3 days with combined aqueous extracts (A) are extracted with hexane to change of media on day 7, allowing for the establishment of yield hexane extract (C) and hydroalcoholic fraction (B). a confluent layer of keratinocytes on the Surface of the gel. The latter is then concentrated until residual water and On day 8, media were removed and culture medium con diluted with ethanol (20%) before extracted with ethyl taining the test plant extracts was added to the wells, acetate to yield aqueous (E) and ethyl acetate extracts (F). followed by 6 or 24 hour incubations at 37° C. in a 5% CO, 0265 All the extracts (A-F) are sampled to verify the controlled atmosphere. The gels were then removed from the process recovery and the aliquots are Submitted to a bio wells and extracted with PBS, with 3 freeze-thaw cycles, followed by centrifugation. The proteolytic activity in the logical evaluation (selective enzymatic inhibition). The Supernatants was assayed by means of a fluorometric assay results are compared with those obtained on the analytical as described above (Example II). scale section and the selected positive extract is then con centrated to dryness under reduced pressure. 0268. The results are provided in Table 6.

TABLE 6

Inhibition of Proteases in a Human Skin Model

Part of % Inhib. 96 Inhib. Plant Stress' plant Concentration Protease 6 hr 24 hr Aconium napeius G 2X MMP-3 O 31 Acorus calamus G 2X MMP-3 O 9 Agrostis alba A. 1X MMP-1 O O Aichemia nois A. O.8X MMP-3 55 41 Allium cepa N FI 2X MMP-2 49 O Aium sativum A. 2X MMP-2 NA 10 Aium Tiberosum A. 1X MMP-3 O 35 Aloe Vera G 2X MMP-2 O O Ambrosia artemisiifolia N LiStFI 2X MMP-9 11 25 Anethlin graveolens A. FIFL St 2X MMP-2 O O Anethlin graveolens G 1X MMP-3 2 31 Anethlin graveolens G 1X MMP-3 O O Anthemistinctoria A. LiSt 2X MMP-3 O 35 Aronia melanocarpa N 2X MMP-3 O 38 (Michx.) Ell. Aronia melanocarpa G 1X MMP-3 O 34 (Michx.) Ell. Aronia Xpriinifolia N LiSt 2X MMP-9 O O Artemisia dractinctius G LiSt 2X MMP-9 O O Artemisia dractinius N LiSt Fr 2X MMP-9 O O Avena Saiiva N 2X MMP-2 O 21 Beta vulgaris G 2X MMP-2 12 10 Beta vulgaris spp. N 2X MMP-2 O O Maritina US 2007/01 22492 A1 May 31, 2007 62

TABLE 6-continued

Inhibition of Proteases in a Human Skin Model

Part of % Inhib. % Inhib. Plant e plant Concentration Protease 6 hr 24 hr

Beta vulgaris Subsp. 2X MM C -2 O O Vulgaris Borago officinalis B 1X MM C 16 O Brassica naptis MM C O O Brassica oleracea MM C NA. 17 Brassica oleracea MM C O Brassica oleracea 0.7X MM C 14 Brassica oleracea FI 1X MM C O Brassica oleracea MM C 16 Brassica rapa 2X MM C O Brassica rapa 2X MM C 10 Bromits inermis 2X MM C O Capsicum annittin Fr 1X MM C 14 Cerasitim to mentostin St 2X MM C 40 Chaerophyllum FI Fr 2X MM C 79 buibosum Chenopodium quinoa St 2X MM C 35 Chichorium endivia 2X MM C 23 Circium arvense St 2X MM C 9 Citritius ianatus MM C O Cornus Canadensis MM C 44 Cynara cardhinctiitis MM C 5 Subsp. Cardinctius Daiiciis Caroia 2X MM C O Daiiciis Caroia 2X MM C O Daiiciis Caroia 2X MM C 12 Dioscorea batatas LF,Fr. 2X MM C O Doichos iabiab FI Fr 2X MM C 1 23 Fagopyrim esculenium 2X MM C O Fagopyri in tatarictim 1X MM C 38 Foeniculum vulgare FI 2X MM C 2O Foeniculum vulgare O.8X MM C 10 Fragaria X ananassa 2X MM C O Frangula aint is Fr 2X MM C 44 Gainsoga St Fl 2X MM C O quadriradiata Glycine max Fr 0.7X Glycyrrhiza glabra 2X MM C Glycyrrhiza glabra St 2X MM C Hamamelis virginiana St 2X MM C 4 Heianthus strumosus 2X MM C Heliotropium FI 2X MM C arborescens Hordeum vulgare 1X MM C -1 1 3 O Subsp. Vulgare Hypomyces lactifiutorum Fr 1X MM C Juniperus Communis FrL St 2X MM C Kochia scoparia St Fr 2X MM C Lactica Saiiva 2X MM C Lentinus edodes Fr 2X MM C 2 1 Lotus corniculatus FrL St 2X MM C Lotus corniculatus C 2X MM C Manihot escuienia Fr O.SX MM C Matricaria rectitita FIFL St O.SX MM C Meilotus albus St 2X MM C Melissa officinalis St O43X MM C Mentha X piperita St Fl 2X MM C 1 Origantin majorana St Fl 2X MM C Panax quinquefoils Fr 2X MM C Pastinaca Saiiva MM C 32 2 Petroselinum crispum MM C Phaiaris canariensis MM C Phaseolus vulgaris O.SX MM C Phaseolus vulgaris O.SX MM C O Physalis philadelphica O.6X MM C 2 Phytolacca decandra F. L. 2X MM C Phytolacca decandra FIFL 2X MM C syn. P. americana Pimpinella anisum FrL St 2X MM C Potentiiia anserina 2X MM C Poterium sanguisorba LS 2X MM C s US 2007/01 22492 A1 May 31, 2007 63

TABLE 6-continued

Inhibition of Proteases in a Human Skin Model

Part of % Inhib. % Inhib. Plant Stress' plant Concentration Protease 6 hr 24 hr Poterium sanguisorba A. LS 2X MMP-3 33 Pyrus communis N Fr 2X MMP-2 41 Raphanus raphanistrum G 0.7X MMP-9 O Rheum rhabarbarium A. 2X MMP-9 36 Ribes nigrum L. A. Fr O.SX MMP-1 24 Ribes Sylvestre N 2X MMP-9 27 Ribes Sylvestre G St 2X MMP-3 33 Rosmarinus officinalis A. LS 2X MMP-3 39 Rubus occidentais N Fr 2X MMP-9 2 14 Rumex crisputs A. R 2X MMP-9 43 Rumex crisputs G R 2X MMP-9 10 Rumex Scittatus N O.SX MMP-9 O Rita graveolens A. LF 1X MMP-3 71 Salvia officinalis A. LS 2X MMP-9 46 Salvia officinalis G St 2X MMP-1 2O Salvia officinalis G St 2X MMP-1 N6 O Sambucus canadensis L. N LFr 2X MMP-2 Saponaria officinalis L. G St 2X MMP-2 Setaria itaica A. LF 2X MMP-2 Soianum melongens N O.SX MMP-1 Soianum melongens N 2X MMP-1 1 Sorghum dochna N 2X MMP-2 bicolor gr technicum Steiaria media N St Fl 2X MMP-2 Steiaria media G St Fl 2X MMP-2 Tanaceilin G 2X MMP-9 o cinerariifolium Taraxacum officinale N 2X MMP-2 2 4o Taraxacum officinale G 2X MMP-2 Teucrium chamaedrys A. St 2X MMP-1 2 Thymus fragantissimus N LS 2X MMP-2 Thymus fragantissimus N LS 2X MMP-2 Thymus praecox. Subsp. A R 1X MMP-1 Arcticus Thymus X citriiodorus G St 2X MMP-2 Trifolium incarnatum N 2X MMP-2 Tropaeoluim maints G FI 2X MMP-2 1 Tropaeoluim maints G 2X MMP-9 1 2 Tropaeoluim maints N O.S6X MMP-9 Tsiiga diversifolia N St 2X MMP-9 Vaccinium N Fr 2X MMP-9 angustifolium Vaccinum angustifolium G St 2X MMP-3 Vitia sp. A. 1X MMP-1 3 Vitia sp. N 1X MMP-3 i X Triticosecale spp. N E 2X MMP-2 Zea mayS G 2X MMP-2 Zea mayS A. LF 1X MMP-2 Zea mayS A. LF 1X MMP-2 Zea mayS G 2X MMP-2 Zea mayS A. LF O.SX MMP-1 O Zea mayS A. LF 2X MMP-2 4 Zea mayS A. LF 2X MMP-2 Zea mayS A. LF 2X MMP-2 Zea mayS N O.SX MMP-9 Zingiber officinale N FrL St 2X MMP-9 'Stress: A: Arachidonic acid; G: Gamma-linolenic acid; N: No stress treatment Part of Plant: B: Buds: E: Ears: Fl: Flower; Fr. Fruit: L: Leaf, R. Root; S: Seed: St: Stem Original screening dose: 1 X = dose at which an inhibition of 50% was obtained in initial screen Ing.

Example VI priate for product development. Stability is ascertained by recovery of protease inhibition over time under various Effect of Plant Extracts on Cell Migration conditions, including physiological conditions. Cytotoxicity is ascertained by incubation of the extracts with various cell 0269 Aqueous and alcoholic plant extracts that inhibit types, including those indicated below. MMP-9, MMP-2 or MMP-1 were prepared as described in Example I and underwent further testing to ascertain that 0270. In order to test the effect of various plant extracts they contain stable, non-cytotoxic molecules that are appro that are also validated protease inhibitors on cellular migra US 2007/01 22492 A1 May 31, 2007 64 tion, a cellular migration assay coupled with a cord forma membrane were wiped off, three randomly selected fields tion assay using endothelial cells was conducted. The were counted on the bottom side. experimental details are provided below. Concentrations of 0274 The percent inhibition of migration is calculated as plant extracts are expressed as a function of the ICso follows: concentration determined for protease inhibition, which is termed IX. The extracts are, therefore, capable of decreasing (A-B)/Ax100, the activity of at least one extracellular protease by at least where A is the average number of cells per field in the 50% when measured according to one of the assays control well and B is the average number of cells per field described herein. The 1x concentration can vary depending in the treated wells. on the plant and the solvent used in the preparation of the 0275 Cord Formation Assay extract. The average concentration of a IX aqueous extract 0276 Matrigel (60 ul of 10 mg/ml) was added to a is about 1.6 mg/ml, whereas the average concentration of a 96-well plate flat bottom plate (Costar 3096) and incubated 1x alcoholic extract is about 4 mg/ml. For each extract tested for 30 minutes at 37°C. in a 5% CO, atmosphere. A mixture in the assays described below, 4 different concentrations of HUVECs and plant extract, or positive controls (Fumag were used (0.31x, 0.62x, 1.25x and 2.5x) in duplicate. illin and GM6001) were added to each well. HUVECs were 0271 Cell Migration Assays prepared as suspensions of 2.5x10 cells per ml in EGM-2, 0272 Migration was assessed using a multi-well system then 500 ul of HUVECs preparation was mixed with 500 ul (Falcon 1185, 24-well format), separated by a PET mem of 2x of the desired dilution of plant extract or control drug brane (8 um pore size) into top and bottom sections. and 200 ul were added to each well. Four dilutions of each Depending on the cells that are used in the assay, the extract were tested in duplicate. After 18-24 hours at 37° C. membrane was coated with 10 ug/ml rat tail collagen and in 5% CO, the cells had migrated and organized into cords allowed to dry. All solutions used in top sections were (see FIG. 4, which shows the results using an extract from prepared in DMEM-0.1% BSA, whereas all solutions used Rheum rhabarbaram). in the bottom sections were DMEM, or other media, con 0277. The number of cell junctions were counted in 3 taining 10% fetal calf serum. randomly selected fields and the inhibition of cord formation is calculated as follows: 0273 EGM-2 (700 ul) was added to the bottom chamber as a chemo-attractant. HUVECs (100 ul of 10° cells/ml) and (A-B)/Ax100, buffer containing the plant extract at the appropriate dilution where A is the average number of cell junctions per field in were added to the upper chamber (duplicate wells of each the control well and B is the average number of cell plant extract at each dilution). After 5 h incubation at 37° C. junctions per field in the treated wells. in a 5% CO, atmosphere, the membrane was rinsed with 0278. The results of the above tests are set forth in Table PBS, fixed and stained. The cells on the upper side of the 7.

TABLE 7 Effect of Exemplary Plant Extracts on Endothelial Cell Migration Endothelial Cell Migration Cellular Migration Assay Cord Formation Assay % inhibition % inhibition Part of Concentration Concentration

Plant Stress' Plant 2.5x 1.25x 0.62x 0.31x 2.5x 1.25x 0.62x 0.31x Allium cepa N 19 28 25 36 O O O O Allium sativum A 16 27 26 34 O O O O Ambrosia artemisiifolia N LiSt 100 90 4 O 99 91 61 57 Ambrosia artemisiifolia N FIFL St 8 S NA NA ND ND ND ND Aronia Xpriinifolia N LiSt 50 26 20 19 ND 93 75 93 Artemisia dractinctius G LiSt 81 57 40 30 45 13 22 23 Artemisia dractinctius N FIFL St 83 SO 41 21 O 6 3 2 Avena Saiiva N 92 75 34 40 1OO 8 O O Beta vulgaris N 30 43 50 47 O O O O Beta vulgaris A O O O O ND ND ND ND Beta vulgaris G 1OO 100 26 SO ND ND ND ND Brassica naptis N ND IND IND IND IND IND IND IND Brassica oleracea N 50 29 30 2O 35 15 O 4 Brassica oleracea N 37 58 23 4 O O O O Brassica oleracea A 65 32 15 21 49 28 27 6 Brassica oleracea A 26 17 O O O O O O Brassica rapa A O 19 31 23 O O O O Brassica rapa N 25 21 14 6 ND ND ND ND Bronts inermis A 90 44 36 17 21 14 O 93 Chenopodium quinoa N LiSt 1OO 100 44 26 90 85 53 42 Chenopodium quinoa N FrL, St 1OO 100 SO 33 ND IND IND IND Subsp. Putinoa US 2007/01 22492 A1 May 31, 2007 65

TABLE 7-continued Effect of Exemplary Plant Extracts on Endothelial Cell Migration Endothelial Cell Migration Cellular Migration Assay Cord Formation Assay % inhibition % inhibition Part of Concentration Concentration

Plant Stress' Plant? 2.5x 1.25x O.62x O.31x 2.5x 1.25x 0.62x 0.31x

Chichorium endivia 83 82 15 O O O O O Chichorium endivia 48 11 21 16 ND IND IND IND Subsp. Endivia Citritius ianatus 88 35 23 14 21 17 6 O Daiiciis Caroia 1OO 63 74 32 92 28 O O Daiiciis Caroia 62 10 O O 53 O O O Daiiciis Caroia O O O O 86 43 25 36 Doichos iabiab FI Fr 60 64 68 83 O O O O Foeniculum vulgare 64 47 62 61 69 21 23 11 Foeniculum vulgare 46 2 34 45 ND IND IND IND Glycyrrhiza glabra 1OO 56 O 53 O O O O Glycyrrhiza glabra St 1OO 34 41 51 O O O O Heianthus strumosus 19 27 2 O 87 68 6 O Hypomyces lactifiutorum Fr 46 30 25 2O 17 O O O Hypomyces lactifiutorum Fr 85 59 31 5 77 67 2O 11 Lentinus edodes Fr 40 16 22 14 O O O O Lotus corniculatus Fr 93 83 77 57 9 O O O Lotus corniculatus C 58 11 26 O O O O O Lotus corniculatus FrL St 18 8 NA NA ND IND IND IND Lotus corniculatus FIFL St 31 3S NA NA ND IND IND IND Lotus corniculatus FIL St 32 36 NA NA ND ND ND ND Manihot escuienia Fr 33 30 25 26 39 O O O Manihot escuienia Fr 69 24 22 31 O 7 O 2O Matricaria rectitita FIFL St 55 45 30 24 O O O O Matricaria rectitita FIFL St 74 6 1 2O 34 3 4 O Meilotus albus St 70 15 O O O O O O Melissa officinalis St 7 10 9 7 ND IND IND IND Phaseolus vulgaris S4 29 10 18 51 17 4 7 Phaseolus vulgaris 82 56 51 41 33 13 25 18 Physalis Philadelphica 1OO 100 100 1OO 100 72 1OO 81 Pimpinella anisum FrL St 70 64 65 69 40 5 27 42 Pistin Saivitin St 38 16 13 O 16 24 4 O Raphantis raphanistrain 88 46 23 23 46 24 O O Raphantis raphanistrain Fr ND IND IND ND ND IND IND IND Rheum X hybridum 13 O NA NA ND IND IND IND (=Rheum rhabarbarum) Ribes Sylvestre 59 49 69 56 96 87 56 26 Rubus occidentais Fr 16 9 O O O O 32 O Rumex crisputs 1OO 86 36 36 95 82 53 48 Rumex crisputs 1OO 11 NA NA ND IND IND IND Rumex Scittatus 1OO 2O O 70 6 O O Setaria itaica LF 93 65 S4 O O O O Sorghum dochna bicolor 32 O O O O O O gr technictim Steiaria media FIFL St 33 27 2 28 O O O O Tanaceilin N 18 21 NA NA ND IND IND IND cinerariifolium Taraxacum officinale 45 11 5 2 O 2 Taraxacum officinale 90 40 44 23 O O O O Thymus fragantissimils St 38 15 1 O O O 22 Thymus X citriiodorus St 76 12 8 32 35 O O Trifolium incarnatum 47 27 5 10 Trifolium incarnatum B.L.St 1OO 100 4 21 ND IND IND IND Tropaeoluim maints 57 58 49 42 O O O O Tropaeoluim maints 65 29 18 Tsiiga Canadensis St 68 41 3 31 ND 8O 82 64 Tsiiga Canadensis St 32 18 NA NA ND IND IND IND Tsiiga diversifolia St 99 43 18 27 57 8 O O Vaccinium anglisiifolium Fr 62 7 24 59 15 6 O X Triticosecale spp. E 8O 84 59 49 O O O O Zea mayS 51 27 O 6 26 25 30 US 2007/01 22492 A1 May 31, 2007 66

TABLE 7-continued Effect of Exemplary Plant Extracts on Endothelial Cell Migration Endothelial Cell Migration Cellular Migration Assay Cord Formation Assay % inhibition % inhibition Part of Concentration Concentration

Plant Stress' Plant 2.5x 1.25x O.62x O.31x 2.5.x 1.25x O.62x 0.31x

Zea mayS A. LF 17 O 49 29 O 6 3 2 Zea mayS N L 66 24 14 6 11 O O 11 Zingiber officinale N Fr 59 38 27 30 O O O O Zingiber officinale N R O 19 NA NA ND ND ND ND 'Stress: A: Arachidonic Acid; G: Gamma-Linolenic Acid; N: No stress treatment Part of Plant: B: Buds: Fl: Flower; Fr. Fruit: L: Leaf; P: Pods: R: Root; S: Seed: St: Stem Original screening dose: 1 X = dose at which an inhibition of 50% was obtained in initial screening.

Example VII sufficient amount of solvent was added to allow moderate agitation with a stirring bar. The solvents used in this Plant Extracts that Inhibit Human Leukocyte Example were: butylene glycol (100%), butylene glycol/ Elastase (HLE) water (50/50, V/v), butylene glycol/water (20/80, V/v); etha 0279 Plant extracts were prepared as described in nol (100%), ethanol/water (85/15, v/v), ethanol/water (50/ Example I and were tested for their ability to inhibit HLE as 50, v/v); water (100%). described in Example II. 0283. Several different extraction times were employed 0280 Results are presented in Table 8. for each solvent: after mixing for periods of 1 hour, 2 hours, 3 hours, or 4 hours at room temperature, the Suspension was TABLE 8 centrifuged and filtered through a 0.45 micron paper filter. For the centrifuged and filtered butylene glycol mixtures, the Inhibition of HLE solvent was then evaporated at 120° C. and the residual Plant Stress Part of Plant matter was weighed to determine the yield of extraction at Arctostaphylos tiva-lisi N LiSt each time point. For the centrifuged and filtered ethanol Arctostaphylos tiva-lisi N LiSt mixtures, the Solvent was removed under reduced pressure Beta vulgaris N R at a temperature of less than 45° C. in order to determine the Corntis sericea G Daiiciis Caroia G yield of extraction at each time point. Euphorbia amygdaloides G LiSt Gainsoga quadriradiata A. Fl 0284. In order to determine the enzymatic and biological Gentiana littea A. properties of the extracts, the 4 hour butylene glycol or Geranium sanguinet in N LiSt ethanol mixtures were assayed without further treatment. Oenothera biennis A. FIFFrLSt Potentilla fruticosa N FIFFrLSt Rodgersia spp. A. 0285) The above protocol is suitable for the preparation Rubus thibetanus G LiSt of extracts that are to be employed in dermatological for Rumex crisputs A. LiFr mulations. Butylene glycol extracts, for example, can be Rumex crisputs G Rumex crisputs N LiFr included directly into formulations intended for topical Vitia sp. A. Fr application. Ethanol extracts may undergo one or more additional steps prior to incorporation into formulations 'Stress: A: Arachidonic Acid; G: Gamma-Linolenic Acid; N: No stress intended for topical application as described in Example XV. treatinent Part of Plant: Fl: Flower; Fr. Fruit: L: Leaf R: Root; S: Seed: St: Stem Example IX Example VIII Protease Inhibition of Plant Extracts Prepared by Preparation of Plant Extracts (Method C) Method C 0281. The following protocol was employed to prepare 0286 Plant extracts prepared as described in Example the plant extracts tested in the following Examples (1x to VIII were tested for their ability to inhibit MMP-1, MMP-2, XIV). MMP-3, MMP-9 and/or HLE using the assays described above (Example II). 0282 For each of the plants, five grams of the dried plant material to be extracted was placed in a beaker and a 0287. The results are presented in Table 9. US 2007/01 22492 A1 May 31, 2007 67

TABLE 9 Inhibition of Proteases by Plant Extracts Prepared by Method C IC50 Plant Part (treatment) Extraction Solvent Enzyme (g/mL) 00% Butylene Glycol MMP-3 NA 1.4 Potentiiia anserina L. 50% Butylene Glycol MMP-3 3O.O 19.0 Aerial parts (untreated) 20% Butylene Glycol MMP-3 92.5 2O.O 00% Ethanol MMP-3 28.8 19.1 Potentiiia anserina L. 85% Ethanol MMP-3 27.6 27.2 Aerial parts (untreated) 50% Ethanol MMP-3 56.3 34.2 OO%. Water MMP-3 58.8 25.7 00% Butylene Glycol MMP-3 35.9 7.7 Rhus typhina L. 50% Butylene Glycol MMP-3 128.6 23.8 Leaf (untreated) 20% Butylene Glycol MMP-3 27.1 22.1 00% Ethanol MMP-3 13.3 9.9 Rhus typhina L. 85% Ethanol MMP-3 27.5 33.4 Leaf (untreated) 50% Ethanol MMP-3 38.0 38.1 OO%. Water MMP-3 54.8 29.0 00% Butylene Glycol MMP-3 42.5 5.8 Juniperus Communis L. 50% Butylene Glycol MMP-3 46.2 14.1 Aerial parts (untreated) 20% Butylene Glycol MMP-3 37.0 12.3 00% Ethanol MMP-3 28 17.5 Juniperus Communis L. 85% Ethanol MMP-3 52 24.O Aerial parts (untreated) 50% Ethanol MMP-3 26 23.9 OO%. Water MMP-3 136 17.1 00% Butylene Glycol MMP-9 19.7 Vaccinium 50% Butylene Glycol MMP-9 58.8 1.8 angustifolium Ait. Press-cake (untreated) 20% Butylene Glycol MMP-9 110.0 1.3 00% Ethanol MMP-9 28.4 7.3 Vaccinium 85% Ethanol MMP-9 29O.S 5.5 angustifolium Ait. Press-cake (untreated) 50% Ethanol MMP-9 11.3 4.0 OO%. Water MMP-9 11.5 2.1 00% Butylene Glycol MMP-9 28.1 1.2 Tropaeoluim maius L. 50% Butylene Glycol MMP-9 108.1 18.3 Aerial parts (G) 20% Butylene Glycol MMP-9 9 O.S 22.9 00% Ethanol MMP-9 48.3 5.2 Tropaeoluim maius L. 85% Ethanol MMP-9 69.0 20.6 Aerial parts (G) 50% Ethanol MMP-9 64.O 33.9 OO%. Water MMP-9 32.9 37.3 00% Butylene Glycol MMP-9 93.0 2.4 Meilotus aiba Medik. 50% Butylene Glycol MMP-9 30.1 13.7 Aerial parts minus main 20% Butylene Glycol MMP-9 30.4 12.6 stem (untreated) 00% Ethanol MMP-9 16.7 6.9 Meilotus aiba Medik. 85% Ethanol MMP-9 19.4 14.8 Aerial parts (untreated) 50% Ethanol MMP-9 60.3 26.5 OO%. Water MMP-9 22.5 28.7 00% Butylene Glycol HL 11.5 1.9 Daiict is carota Subsp. 50% Butylene Glycol HL 12.2 15.2 Carota L. Aerial parts (untreated) 20% Butylene Glycol HL 68.3 16.O 00% Ethanol HLE 2O2 4.7 Daiict is carota Subsp. 85% Ethanol HLE 8.3 12.3 Carota L. Aerial parts (untreated) 50% Ethanol l 5.8 22.6 OO%. Water l 43.1 21.6 00% Butylene Glycol HL O.35 O.O Geranium x cantabrigiense 50% Butylene Glycol HL 14.10 8.3 Leaf (untreated) 20% Butylene Glycol HL 1140 7.0 00% Ethanol l O.31 5.7 Geranium x cantabrigiense 85% Ethanol l 0.27 15.9 Leaf (untreated) 50% Ethanol l O.35 29.9 OO%. Water l O.43 21.5 00% Butylene Glycol MMP-9 16.5 6.6 Chenopodium quinoa 50% Butylene Glycol MMP-9 S.6 10.6 Willd. (Norquin) Seed (untreated) 20% Butylene Glycol MMP-9 5.4 6.3 00% Ethanol MMP-9 20.4 7.0 Chenopodium quinoa 85% Ethanol MMP-9 13.4 5.8 Willd. (Norquin) Seed (untreated) 50% Ethanol MMP-9 13.8 6.8 OO%. Water MMP-9 6.8 11.2 00% Butylene Glycol MMP-2 O.35 O.O US 2007/01 22492 A1 May 31, 2007 68

TABLE 9-continued Inhibition of Proteases by Plant Extracts Prepared by Method C IC50 Plant Part (treatment) Extraction Solvent Enzyme (g/mL) a Triticosecale spp. 50% Butylene Glycol MM P-2 14.10 8.3 Seed (untreated) 20% Butylene Glycol MM P-2 1140 7.0 00% Ethanol MM P-2 11.0 2.2 a Triticosecale spp. 85% Ethanol MM P-2 2.4 4.4 Seed (untreated) 50% Ethanol MM P-2 3.3 9.2 OO%. Water MM P-2 3.7 10.4 00% Butylene Glycol MM P-9 7.5 O.8 Chenopodium quinoa 50% Butylene Glycol MM P-9 98 6.1 Willd. (Royal) Seed (untreated) 20% Butylene Glycol MM 58.3 7.3 00% Ethanol MM 16.3 7.4 Chenopodium quinoa 8 50% Ethanol MM 8.4 S.O Willd. (Royal) Seed (untreated) 5 09% Ethanol MM 19.0 5.8 OO%. Water MM 2.8 10.8 00% Butylene Glycol MM 17 5.5 Beta vulgaris L. Subsp. 50% Butylene Glycol MM 17.8 22.8 Vulgaris Leaf (sandblasted) 20% Butylene Glycol MM 26.1 18.8 00% Ethanol MM 13.5 7.8 85% Ethanol MM 62 18.2 Beta vulgaris L. Subsp. 50% Ethanol MM 45 23.7 Vulgaris Leaf (sandblasted) OO%. Water MM 8.2 25.3 00% Butylene Glycol MM 40 1.9 Zea mays L. 50% Butylene Glycol MM 25 14.1 Leaf (untreated) 20% Butylene Glycol MM 2O 14.1 00% Ethanol MM 2S6 S.O 85% Ethanol MM 338 8.O Zea mays L. 50% Ethanol MM 40S 12.8 Leaf (untreated) OO%. Water MM 146 14.3 00% Butylene Glycol MM 35 1.9 Zea mays L. 50% Butylene Glycol MM 14.1 Leaf (untreated) 20% Butylene Glycol MM 14.1 00% Butylene Glycol MM 140 3.9 Brassica oleracea L. 50% Butylene Glycol MM 117 20.7 war. iiaica Pienck Head (untreated) 20% Butylene Glycol MM 78 23.1 00% Ethanol MM 31.5 6.2 85% Ethanol MM 146S 24.7 Brassica oleracea L. 50% Ethanol MM Negative 33.5 war. iiaica Pienck Head (untreated) OO%. Water MM 105 29.0 00% Butylene Glycol MM 8O 1.6 Capsicum annittin L. 50% Butylene Glycol MM 140 23.5 W8. (iiitiii. Leaf (untreated) 20% Butylene Glycol MM 112 22.4 00% Ethanol MM 323 6.6 85% Ethanol MM 760 22.7 Capsicum annittin L. 50% Ethanol MM 788 36.5 W8. (iiitiii. Leaf (untreated) OO%. Water MM 57 26.O 00% Butylene Glycol MM 35 3.8 Soianum melongena L. 50% Butylene Glycol MM 1100 22.2 Leaf (untreated) 20% Butylene Glycol MM 805 24.9 00% Ethanol MM 88 12.8 85% Ethanol MM 96.O 38.3 50% Ethanol MM Negative 37.5 OO%. Water MM 654 38.8 00% Butylene Glycol MM 23 2.9 50% Butylene Glycol MM 2O1 24.6 Root (untreated) 20% Butylene Glycol MM 140 25.4 00% Ethanol MM 53 5.8 hanol MM 204 19.0 Pastinaca Saiiva L. hanol MM 365 27.1 Root (untreated) MM 459 28.4 US 2007/01 22492 A1 May 31, 2007 69

Example X 5x10 cells/100 ul/well and keratinocytes at 8x10 cells/100 ul well. Plates were incubated for 24 hours at 37° C. in a Cytotoxicity Testing of Plant Extracts humidified 5% CO atmosphere. The extracts were obtained 0288 This example describes a method of testing the and diluted at a concentration 2 mg/ml (2x the final con plant extracts for their cytotoxicity and allows non-cytotoxic centration) in both media. Four dilutions were tested for concentrations of the extracts suitable for further efficacy each cell line. Controls were included for each assay, 100 ul studies to be selected. Plant extracts were prepared as of media to reflect the maximum growth and viability of described in Example VIII. cells and 100 ng/ml of daunorubicin to obtain an 80% cytotoxic effect. All wells were incubated for 72 hours at 37° 0289 Normal human skin fibroblasts and keratinocytes C. in a humidified 5% CO atmosphere. After incubation, (Cascade Biologics, Portland, Oreg.) were tested to evaluate Alamar Blue dye was added to each well', fluorescence was the possible anti-proliferative effect of an extract of the read on a Spectrafluor Plus (Tecan, Durham, N.C.). All present invention. The latter was done to ascertain that the assays were done in quadruplicate. exposure of cells to a concentration of extract would have no undesirable effect for further cellular assays. The present 0290 The results are shown in Table 10. FIG. 7 presents response was measured in a 96-well plate. Cells were seeded the results for the extracts from Melilotus alba and Junipe in their media (M106+LSGS for fibroblasts and M154+ rus communis. The results represent the average of quadru HKGS for keratinocytes, Cascade Biologics) fibroblasts at plicate measurements.

TABLE 10 Cytotoxicity of Representative Plant Extracts IC50/100% viability Extraction in mg ml Plant Plant part Solvent (yfy) Protease Keratinocytes Fibroblasts Potentia Aerial parts BG’?water 50:50 MMP-3 0.120.1 O.3S.O.3 anSerina L. BG/water 20:80 MMP-3 0.04f0.02 O.7.0.3 Rhus typhina L Leaf BG/water 50:50 MMP-3 &O.O3 O.S.O.1 BG/water 20:80 MMP-3 &O.O3 0.40.1 Juniperus Aerial parts BG/water 50:50 MMP-3 O.07.0.03 O3,O.O3 communis L. BG/water 20:80 MMP-3 O.33,0.12 1.O.25 Tropaeoliin Aerial parts BG/water 50:50 MMP-9 100% viable at 100% viable at maius L. O.6 O6 BG/water 20:80 MMP-9 100% viable at 100% viable at O.8 O.9 Meilotus aiba Aerial parts BG/water 50:50 MMP-9 100% viable at 1 100% viable at 1 Medik. (minus main BG/water 20:80 MMP-9 100% viable at 1 100% viable at 1 stem) Daiiciis Caroia Aerial parts BG/water 50:50 HLE 0.2.0.1 O.S.S.O.3 Subsp carota L. BG/water 20:80 HLE 1.O.3 100% viable at O.1 Geranium X catabrigiense Leaf BG/water 100:0 HLE O.O2S.O.O17 O.O2S.O.O17 BG/water 50:50 HLE O.17.O.O33 O.6,0.33 BG/water 20:80 HLE 0.1.0.03 10.6 Beta vulgaris L. Leaf BG/water 50:50 MMP-2 00% viable at 100% viable at 1 Subsp. Vulgaris 0.7 BG/water 20:80 MMP- 100% viable at 1 100% viable at 1 Zea mays L. Leaf BG/water 50:50 MMP- 00% viable a 100% viable at et-9 O.6 O.2 BG/water 20:80 MMP-1 100% viable at 1 100% viable at et-9 O.3 Brassica Head BG/water 50:50 MMP-1 00% viable a O.S.S.O. oleracea L. var. 0.7 itaica Pienck BG/water 20:80 MMP-1 00% viable a O.6S.O.3 O.8 Chenopodium Seed BG/water 0:100 MMP-9 00% viable at 100% viable at 1 quinoa Willd. O.8 Triticosecale Seed EtOH water MMP-2 O.48.O.25 100% viable at spp. 100:0 O6 Pastinaca Saiiva Root BG/water 50:50 MMP-1 00% viable a 100% viable at L. O.8 O.8 BG/water 20:80 MMP-1 100% viable at 1 100% viable at 0.75 Pastinaca Saiiva Root EtOH water MMP-1 O.07.0.04 O.1.O.O7 L. 100:0 BG/water 100:0 MMP-1 0.110.08 100% viable at O.12 BG/water 20:80 MMP-1 100% viable at 100% viable at 1 US 2007/01 22492 A1 May 31, 2007 70

TABLE 10-continued Cytotoxicity of Representative Plant Extracts IC50/100% viability Extraction in mg ml Plant Plant part Solvent (yfy) Protease Keratinocytes Fibroblasts Brassica Head BG/water 100:0 MMP-1 0.04f0.01 O.O7,0.04 oleracea L. var. BG/water 50:50 MMP-1 0.8.0.1 100% viable at itaica Pienck O.8 BG/water 20:80 MMP-1 O.7 O.1 100% viable at 0.4 Capsicum Leaf EtOH water MMP-1 0.1.0.03 O.6,0.35 annittin L. var. 20:80 (iitiitii BG/water O:100 MMP-1 O.2S.O.OS 0.7.0.5 Soianum Leaf EtOH water MMP-1 0.07.0.0.4 O.O7,0.04 melongena 100:0 BG/water 100:0 MMP-1 O.09.O.O3S 0.120.08 This value represents the concentration at which 100% viability is retained in the tested cell line. “BG: butylene glycol

Example XI 0294 The ELISA was performed following the protocol recommended by the manufacturer (Takara Biomedicals). Effect of Plant Extracts on Collagen Production 20 ul of the supernatant from each well were used. Standard 0291. This following example demonstrates the ability of buffer and stop solutions were freshly prepared. 100 ul of the exemplary plant extracts to stimulate collagen I production antibody-POD conjugate was added into the wells of the in human dermal fibroblast cells. Human dermal fibroblast pre-coated 96-well plate, then the 20 ul of standard and cells (Cascade Biologics, Portland, Oreg.) were employed in specimens were added to appropriate wells. The plate was the assay and the ability of the plant extract to stimulate collagen production was measured using the Takara Bio mixed gently, sealed (to limit evaporation) and incubated for medicals ELISA kit (Takara Mirus Bio, Madison, Wis.), 3 hours at 37° C. which evaluates the release of the procollagen type I C-pep tide (PIP). This free propeptide indicates on a stoichiometric 0295) After incubation, each well was washed four times basis the number of collagen molecules synthesised since with PBS buffer. 100 ul of the substrate solution containing the PEP peptide is cleaved off the procollagen molecule hydrogen peroxide and tetramethylbenzidine (TMBZ) was during the formation of the collagen triple helix. Plant added to each well and the plate was incubated for 15 extracts were prepared as described in Example VIII. minutes. After incubation 100 ul of 1N HSO (stop solu 0292 Fibroblasts were first grown in a 96-well plate tion) was added to each well. The plate was then gently using the complete M106 (M106+LSGS: Cascade Biolog mixed and the absorbance was read at 450 nm on the ics). This media was also used as control. GM6001 (Chemi Spectrafluor Plus plate reader (Tecan). The reading was con, Temecula, CA) was used as positive control at a taken within 15 minutes of addition of the stop solution. All concentration of 50 uM. solutions used were included in the kit except for the PBS 0293 All extracts and controls were diluted in complete and the stop solution. M106. Plant extracts were used at the concentration that provided 100% viability of fibroblasts as shown in Table 10. 0296) The results are presented in Table 11. FIG. 8 Cells were seeded into 96-well plates at a concentration of presents results of extracts for various extracts (A: extract 5x10 cells/well in complete M106 media. Plates were using 50:50 V/v butylene glycol: water as solvent; B: extract incubated for 72 hours at 37° C. in a humidified 5% CO, atmosphere. After incubation, the medium was removed and using 20:80 V/v butylene glycol: water as solvent). The 200 ul of sample were added to the wells (all in duplicate). control (Mock BU:HO and cells alone) demonstrated the Plates were incubated for 48 hours at 37° C. in a humidified lowest collagen I production compared to the positive con 5% CO atmosphere. trol GM6001 at 50 uM.

TABLE 11 Increase in PIP Production Stimulated by Representative Plant Extracts

Plant Plant part Extraction Solvent (viv) PIP? (% increase) Potentiiia anserina Aerial parts BG/water 50:50 Negative L. BG/water 20:80 Negative Rhus typhina L Leaf BG/water 50:50 Negative BG/water 20:80 Positive? (+133%) US 2007/01 22492 A1 May 31, 2007 71

TABLE 11-continued Increase in PIP Production Stimulated by Representative Plant Extracts Plant Plant part Extraction Solvent (v/v) PIP (% increase) Juniperus Aerial parts BG/water 50:50 Positive? (+25%) communis L. BG/water 20:80 Positive? (+11.1%) Tropaeoluim maints Aerial parts BG/water 50:50 Positive? (+42%) L. BG/water 20:80 Negative Meilotus aiba Aerial parts BG/water 50:50 Negative Medik. (minus main BG water 20:80 Positive? (+36%) stem) Daiiciis Caroia Aerial parts BG/water 50:50 Negative Subsp carota L. BG/water 20:80 Negative Geranium x catabrigiense Leaf BG/water 100:0 Negative BG/water 50:50 Negative BG/water 20:80 Negative Beta vulgaris L. Leaf BG/water 50:50 Negative Subsp. Vulgaris BG/water 20:80 Negative Zea mays L. Leaf BG/water 50:50 Positive? (+17%) BG/water 20:80 Negative Brassica oleracea Head BG/water 50:50 Positive? (+11%) L. var. italica BG/water 20:80 Positive? (+1.5%) Pienick Chenopodium Seed BG/water 0:100 Positive/(+8%) quinoa Willd. Triticosecale spp. Seed EtOH/water 100:0 Positive? (+21%) Pastinaca Saiiva L. Root BG/water 50:50 Positive? (+14%) BG/water 20:80 Positive? (+11%) Pastinaca Saiiva L. Root EtOH/water 100:0 Negative BG/water 100:0 Positive/(+57.5%) BG/water 20:80) Positive/(+72.5%) Brassica oleracea Head BG/water 100:0 Positive? (+36.0%) L. var. italica BG/water 50:50 Negative Pienick BG/water 20:80 Positive/(+67.9%) Capsicum annittin Leaf EtOH/water 20:80 Positive? (+34.1%) L. war. annittin BG/water 0:100 Positive? (+3.06%) Soianum Leaf EtOH/water 100:0 Negative melongena BG/water 100:0 Positive? (+21.3%) BG: butylene glycol

Example XII of collagen I collagen I (rat tail, BD Biosciences, Bedford, Mass.), M106 5x, NaOH 0.7N; 1x10 cells and fetal bovine Inhibition of Dermal Contraction by Plant Extracts serum (FBS) at 20% (Wisent, St-Bruno, QC, Canada). The 0297. The following example demonstrates the ability of mix was kept on ice until distribution. The derm-like gels exemplary extracts prepared as described in Example VIII to were allowed to polymerize for 1 hour at 37° C. in a inhibit dermal contraction in an in vitro skin model. The skin humidified 5% CO, atmosphere. After incubation, the gels model comprises human skin fibroblasts imbedded in a were detached from the wells. Media 106 was used as collagen I matrix and provides an in vitro representation of negative control and GM6001 (Chemicon, Temecula, CA) at dermal contraction resulting from tractional forces gener a concentration of 50 uM was used as positive control. All ated by fibroblasts. Partial or permanent dermal contraction extracts were prepared at non-cytotoxic concentration (i.e. can play a role in the formation of wrinkles. Thus, extracts the concentration that provided 100% viability of fibroblasts capable of inhibiting this type of contraction, have the as shown in Table 10) in complete media 106. FBS at a final potential to provide a dermo-decontraction anti-ageing concentration of 10% was added to each well. The plate was effect in the skin. These extracts also have potential appli incubated for a maximum of 7 days at 37°C. in a humidified cation in wound healing where pathological scarring is 5% CO atmosphere. All assays were performed in dupli observed by excessive contraction. cate. Contraction was measured beginning at day 3. Con 0298 The ability of exemplary plant extracts to inhibit tracting gels were digitally photographed and the gel areas dermal contraction was evaluated on human skin fibroblasts were calculated using ImagePro Software. (Cascade Biologics, Portland, Oreg.). The cells were imbed ded in a collagen I matrix to create a derm-like environment. 0299) The results are presented in Table 12. Control gels Fibroblasts were grown in complete M106 to 80% conflu treated with media alone have the Smallest area and repre ence. Free-floating fibroblast-populated collagen gels were sent the contracted control. GM6001 was able to provide prepared in 24-well plates. 500 Lulof gel contains 2.5 mg/ml limited, but not complete, inhibition of contraction. US 2007/01 22492 A1 May 31, 2007 72

Oakville, Ontario) at a concentration of 2.5 LM was used as TABLE 12 a positive control. All tested plant extracts and the controls were diluted in complete M154 at a non-cytotoxic concen Inhibition of Dernal Contraction by Representative Plant Extracts tration (i.e. the concentration that provided 100% viability of Contraction keratinocytes as shown in Table 10). Cells were seeded into Extraction Solvent (% 96-well plates at a concentration of 8x10 cells/well in Plant Plant part (v/v) inhibition) complete M154 media. Plates were incubated for 48 hours at 37° C. in a humidified 5% CO, atmosphere. After incu Potentilla anserina L. Aerial parts BG/water 50:50 96.5 BG/water 20:80 94.5 bation, the medium was removed and 200 ul of sample was Rhus typhina L Leaf BG/water 50:50 79.7 added to the wells (all in duplicate). Plates were incubated BG/water 20:80 39.2 for a further 48 hours at 37° C. in a humidified 5% CO, Juniperus Aerial parts BG/water 50:50 86.4 atmosphere. communis L. BG/water 20:80 86.5 Tropaeolum maius L. Aerial parts BG/water 50:50 44.2 0302) The ELISA was performed using following the BG/water 20:80 44.2 protocol recommended by the manufacturer (R&D Sys Melilotus alba Medik. Aerial parts BG/water 50:50 62.8 (minus BG/water 20:80 48.5 tems). 25 ul of the supernatant from each well was mixed main stem) with 25 ul of R5DP 1x diluting buffer. Standards were Beta vulgaris L. Leaf BG/water 50:50 13.3 freshly prepared in R5DP 1x. 100 ul of assay diluent RD1-8 Subsp. Vulgaris BG/water 20:80 41.6 was added to each well of 96-well plate, then the 50 ul of Zea mays L. Leaf BG/water 50:50 22.4 standard and specimens were added to appropriate wells. BG/water 20:80 100 The plate was mixed gently, sealed (to limit evaporation) Brassica oieracea L. Head BG/water 50:50 2O2 war. italica Pienck BG/water 20:80 4.3 and incubated for 2 hours at room temperature (RT). Chenopodium quinoa Seed BG/water 0:100 O Wild. 0303. After incubation, each well was washed four times Triticosecale spp. Seed EtOH/water 100:0 -11 with wash buffer. 100 ul of the conjugation solution was Pastinaca Saiiva L. Root BG/water 50:50 15.7 added and incubated for 1 hour at RT. After this incubation, BG/water 20:80 6.2 each well was washed four times with wash buffer. 200 ul of Pastinaca Saiiva L. Root EtOH/water 100:0 624 Substrate Solution containing hydrogen peroxide and tetram BG/water 100:0 25.7 BG/water 20:80 17.5 ethylbenzidine (TMBZ) was added to each well and the Brassica oieracea L. Head BG/water 100:0 14.7 plate was incubated for 15 minutes. After incubation, 50 ul war. iiaica Pienck BG/water 50:50 6.7 of 2N HSO (stop solution) was added to each well. The BG/water 20:80 21.2 plate was then gently mixed and the absorbance read at 450 Capsicum annittin L. Leaf EtOH/water 20:80 13 nm on the Spectrafluor Plus plate reader (Tecan). The W8. (iiitiii. BG/water 0:100 37.6 Soianum melongena Leaf EtOH/water 100:0 33.4 reading was taken within 15 minutes following addition of BG/water 100:0 7.2 the stop solution. Ali solutions employed were provided in the kit. BG: butylene glycol 0304) The above-described ELISA evaluates the release of IL-8. A plant extracts that results in a strong release of Example XIII IL-8 may cause irritation to the skin at the tested concen tration. The results are shown in Table 13. FIG. 9 presents Effect of Plant Extracts on Cytokine Release results for various extracts (A: extract using 50:50 V/v butylene glycol: water as solvent; B: extract using 20:80 V/v 0300. The following example demonstrates the non-irri butylene glycol: water as solvent; C: extract using 100% tating behaviour of representative plant extracts of the water as solvent; D: extract using 100% ethanol as solvent). invention prepared as described in Example VIII. The The negative control (M154 media) showed the lowest IL-8 amount of Interleukin-8 (IL-8) released after exposure of release and is considered to represent the minimum IL-8 keratinocytes to a plant extract, as described below, can be release. PMA induced a strong inflammatory response and is used to quantify any possible irritation reaction to the considered to represent the highest level of IL-8 release. eXtract. Although some of tested extracts increased in IL-8 release, 0301 IL-8 release was evaluated in human skin kerati the increase was small compared to that induced by PMA. nocytes (Cascade Biologics, Portland, Oreg.) using the Those extracts resulting in a small increase in IL-8 release Quantikine hIL-8 ELISA kit (R&D Systems, Minneapolis, can be re-assayed at a lower concentration, which will likely Minn.). Keratinocytes were first grown in a 96-well plate result in a relative decrease in the amount of IL-8 released. using the complete M154 (M154+HKGS from Cascade The evaluation of cytokine release as described above Biologics). This media was also used as control. Phorbol enables a maximum concentration of plant extract for further 12-myristate 13-acetate (PMA) (Sigma-Aldrich Canada, in vivo studies to be set.

TABLE 13 Effect of Representative Plant Extracts on IL-8 Release Plant Plant part Extraction Solvent (v/v) IL-8 (% change)' Potentiiia anserina Aerial parts BG’/water 50:50 -83 BG/water 20:80 -79 Rhus typhina L Leaf BG/water 50:50 95 BG/water 20:80 -61 US 2007/01 22492 A1 May 31, 2007 73

TABLE 13-continued Effect of Representative Plant Extracts on IL-8 Release Plant Plant part Extraction Solvent (v/v) IL-8 (% change)' Juniperus Aerial parts BG/water 50:50 -83 communis L. BG/water 20:80 -79 Tropaeoluim maints Aerial parts BG/water 50:50 -86 L. BG/water 20:80 -77 Meilotus aiba Aerial parts BG/water 50:50 Performed at 2 Medik (minus main concentrations: stem) At 1 mg: 226 At 0.3 mg: 84 BG/water 20:80 -46 Daiiciis Caroia Aerial parts BG/water 50:50 -74 Subsp carota L. BG/water 20:80 -61 Geranium x catabrigiense Leaf BG/water 100:0 7 BG/water 50:50 -60 BG/water 20:80 -53 Beta vulgaris L. Leaf BG/water 50:50 133 Subsp. Vulgaris BG/water 20:80 Performed at 2 concentrations: At 1 mg: 158 At 0.3 mg: 54 Zea mays L. Leaf BG/water 50:50 -13 BG/water 20:80 -13 Brassica oleracea Head BG/water 50:50 -7 L. var: italica BG/water 20:80 41 Pienick Chenopodium Seed BG/water 0:100 Performed at 2 quinoa Willd. concentrations: At 1 mg: 264 At 0.3 mg: 105 Triticosecale spp. Seed EtOH/water 100:0 3.2 Pastinaca Saiiva L. Root BG/water 50:50 72 BG/water 20:80 190 Pastinaca Saiiva L. Root EtOH/water 100:0 36 BG/water 100:0 103 BG/water 20:80 -67 Brassica oleracea Head BG/water 100:0 -67 L. var. italica BG/water 50:50 Performed at 2 Pienick concentrations: At 1 mg: 201 At 0.3 mg: 159 BG/water 20:80 13 Capsicum annittin Leaf EtOH/water 20:80 -39 L. war. annittin BG/water 0:100 S4 Soianum Leaf EtOH/water 100:0 -65 melongena BG/water 100:0 O Value indicates the 96 change relative to the negative control (untreated cells) °BG: butylene glycol

Example XIV After irradiation, test samples were added at 500 ul/well. The media was used as a negative control. GM6001 at a Inhibition of UV-Induced Proteolytic Activity concentration of 50 uM was used as a positive control. All 0305 The following example demonstrates the potential extracts and controls were diluted in complete M154 at a of representative plant extracts to protect the skin from non-cytotoxic concentration (i.e. the concentration that pro proteolytic damage after Sun exposure. Plant extracts were vided 100% viability of keratinocytes as shown in Table 10). prepared as described in Example VIII. Plates were incubated for 24 hours at 37°C. in a humidified 5% CO atmosphere. The supernatant from each well was 0306 Keratinocytes were first grown in 24-well plates assayed or kept at -80° C. until use. Supernatants (60 ul) using the complete M154 (M154+HKGS from Cascade were assayed for their overall proteolytic activity using the Biologics) at a concentration of 2.5x10" cells/500 ul/well. MMP2/7 internally quenched peptide (Calbiochem). All: The plates were incubated 48 hours at 37°C. in a humidified assays were performed in duplicate except for controls, 5% CO, atmosphere. The media was removed and the cells which were performed in quadruplicate. were washed 2 times with HBSS. After complete removal of liquid the cells were irradiated with 25 J/cm of WVA light. 0307 The results are presented in Table 14. US 2007/01 22492 A1 May 31, 2007 74

TABLE 1.4 Inhibition of UV-Induced Protease Activity by Representative Plant Extracts

Extraction Solvent Decrease in Plant Plant part (v/v) Protease Activity' Potentiiia anserina L. Aerial parts BG’/water 50:50 73.5 BG/water 20:80 O Rhus typhina L Leaf BG/water 50:50 28 BG/water 20:80 16.5 Juniperus Communis L. Aerial parts BG/water 50:50 46.5 BG/water 20:80 15 Tropaeoluim maius L. Aerial parts BG/water 50:50 O BG/water 20:80 Meilotus aiba Medik. Aerial parts (minus BG/water 50:50 main stem) BG/water 20:80 Daiict is carota Subsp. Aerial parts BG/water 50:50 Carota L. BG/water 20:80 Geranium x catabrigiense Leaf BG/water 100:0 BG/water 50:50 BG/water 20:80 4 Beta vulgaris L. Leaf BG/water 50:50 Subsp. Vulgaris BG/water 20:80 Zea mays L. Leaf BG/water 50:50 BG/water 20:80 1 Brassica oleracea L. Head BG/water 50:50 war. iiaica Pienck BG/water 20:80 Chenopodium quinoa Seed BG/water 0:100 Wild. Triticosecale spp. Seed EtOH water 100:0 Pastinaca Saiiva L. Root BG/water 50:50 BG/water 20:80) Pastinaca Saiiva L. Root EtOH water 3 5 100:0 BG/water 100:0 1 BG/water 20:80 Brassica oleracea L. Head BG/water 100:0 war. iiaica Pienck BG/water 50:50 BG/water 20:80 Capsicum annittin L. Leaf EtOH water W8. (iiitiii. 20:80 BG/water 0:100 Soianum melongena Leaf EtOH water g 100:0 BG/water 100:0 O "Decrease in proteolytic activity on MMP 2/7 peptide after UV irradiation relative to untreated control. °BG: butylene glycol

Example XV The activity of the extract may be verified at one or several points in this additional procedure. Preparation of Ethanolic Plant Extracts for Topical 0309 The disclosure of all patents, publications, includ Formulations ing published patent applications, and database entries ref 0308) As ethanol is not commonly used as a solvent in erenced in this specification are specifically incorporated by cosmetic formulations, plant extracts prepared by ethanolic reference in their entirety to the same extent as if each such extractions as described in Example VIII can undergo fur individual patent, publication, and database entry were spe ther treatments to prepare them for incorporation into topical cifically and individually indicated to be incorporated by formulations. For example, the ethanolic extracts can be reference. de-colourised by treatment with activated charcoal follow ing standard protocols. The ethanol can be removed from 0310. The invention being thus described, it will be extracts, or de-colourised extracts and the reduced extract obvious that the same may be varied in many ways. Such material resuspended on a solid Support or in a liquid solvent variations are not to be regarded as a departure from the that is more acceptable to cosmetic formulators. Thus, the spirit and scope of the invention, and all Such modifications extracts, or de-colourised extracts, can be submitted to an as would be obvious to one skilled in the art are intended to evaporation procedure (for example using a rotary evapo be included within the scope of the following claims. rator or soxlet) to remove some, or all, of the ethanol component of the solvent. A dermatologically suitable alco 1. A dermatological formulation comprising a physiologi hol. Such as a glycol, can be added and the resulting Solution cally acceptable carrier and an effective amount of one or incorporated into a carrier Suitable for topical application. more plant extracts having extracellular protease inhibiting US 2007/01 22492 A1 May 31, 2007 activity, said plant extract derived from any one of the plants 20. The process according to claim 18, wherein said listed in Tables 1, 2, 3, 4 and 5 by solvent extraction, and said plurality of potential extracts is generated by selecting a extracellular protease selected from the group of matrix group of plants; harvesting plant material from each plant in metalloprotease-1 (MMP-1), matrix metalloprotease-2 said selected group of plants; and Subjecting said plant (MMP-2), matrix metalloprotease-3 (MMP-3), matrix met material from each plant to a solvent extraction process to alloprotease-9 (MMP-9) and human leukocyte elastase provide said plurality of potential extracts. (HLE), wherein said plant extract modulates one or more cellular activities in skin cells. 21. The process according to claim 18, wherein said 2. The dermatological formulation according to claim 1, Solvent extraction process employs an alcohol, water, an wherein said one or more cellular activities in skin cells are aqueous buffer, or a combination thereof as solvent. selected from the group of attenuating the breakdown of 22. The process according to claim 18, wherein the group collagen, fibronectin, fibrillin and/or elastin; attenuating of extracts selected in step (c) are capable of inhibiting the endothelial cell migration; increasing collagen production; activity of at least one of said extracellular proteases by at attenuating UV-induced extracellular protease activity and least 20%. attenuating tractional forces generated by fibroblasts. 23. The process according to claim 18, further comprising 3. The dermatological formulation according to claim 1, the steps of Subjecting each plant extract in said group of wherein said solvent is an aqueous solvent, an alcoholic extracts to at least one cytotoxicity, bioavailability or sta Solvent, or a combination thereof. bility test and selecting those extracts that demonstrate 4.-9. (canceled) physiologically acceptable cytotoxicity, bioavailability and/ 10. The dermatological formulation according to claim 1, wherein said dermatological formulation is for use in the or stability. routine care of the skin, hair and/or nails. 24. The process according to claim 18, wherein said plant 11. The dermatological formulation according to claim 1, material is generated from a plant or group of plants that wherein said dermatological formulation is for use to have been Subjected to one or more stress. improve the health and/or appearance of the skin, hair and/or 25. The dermatological formulation according to claim 1, nails. wherein said plant extract having extracellular protease 12. The dermatological formulation according to claim 1, inhibiting activity is derived by solvent extraction from a wherein said dermatological formulation is for use in the plant selected from the group of Aconitum napellus, Acorus treatment or prevention of a dermatological condition. Calamus, Alchemilla mollis, Allium cepa, Allium sativum, 13. The dermatological formulation according to claim 1, Allium tuberosum, Ambrosia artemisiifolia, Anethum gra wherein said dermatological formulation is for use to attenu veolens, Anthemis tinctoria, Aronia melanocarpa (Michx.) ate or prevent skin ageing. Ell. Arctostaphylos uva-ursi, Aroniaxprunifolia, Artemisia 14.-17. (canceled) 18. A process for identifying a plant extract suitable for dracunculus, Avena sativa, Beta vulgaris, Beta vulgaris L. the preparation of a dermatological formulation, said pro Subsp. Vulgaris, Borago officinalis, Brassica napus, Bras cess comprising the steps of sica oleracea, Brassica oleracea L. var. italica Plenck, Brassica rapa, Bromus inermis, Capsicum annuum L. var. (a) generating a plurality of potential extracts by solvent annuum, Cerastium tomentosum, Chaerophyllum bulbosum, extraction of plant material; Chenopodium quinoa, Chenopodium quinoa Subsp. Quinoa, (b) analysing the ability of each of said potential plant Chenopodium quinoa Willd. Chichorium endivia, Chicho extracts to inhibit one or more extracellular protease rium endivia Subsp. Endivia, Circium arvense, Citrullus Selected from the group of matrix metalloprotease-1 lanatus, Cornus Canadensis, Cornus sericea, Cynara car dunculus Subsp. Cardunculus, Daucus carota, Daucus (MMP-1), matrix metalloprotease-2 (MMP-2), matrix carota Subsp carota L., Dolichos lablab, Euphorbia metalloprotease-3 (MMP-3), matrix metalloprotease-9 amygdaloides, Fagopyrum tataricum, Foeniculum vulgare, (MMP-9) and human leukocyte elastase (HLE); Frangula alnus, Galinsoga quadriradiata, Gentiana lutea, (c) selecting those potential extracts that are capable of Geranium sanguineum, Geraniumxcantabrigiense, Glycyr inhibiting the activity of at least one of said extracel rhiza glabra, Hamamelis virginiana, Helianthus strumosus, lular proteases to provide a group of extracts; Heliotropium arborescens, Hordeum vulgare subsp. Vul (d) analysing each extract in said group of extracts for the gare, Hypomyces lactifluorum, Juniperus communis L., ability to modulate one or more cellular activities in Lentinus edodes, Lotus corniculatus, Manihot esculenta, skin cells selected from the group of attenuating the Matricaria recutita, Melilotus albus, Melilotus alba Medik., breakdown of collagen, fibronectin, fibrillin and/or Melissa officinalis, Menthaxpiperita, Oenothera biennis, elastin, attenuating endothelial cell migration; increas Pastinaca sativa L., Petroselinum crispum, Phaseolus vul ing collagen production; attenuating UV-induced extra garis, Physalis philadelphica, Phytolacca decandra, Phyto cellular protease activity and attenuating tractional lacca decandra syn. P. americana, Pimpinella anisum, Pisum sativum, Potentilla anserina L., Potentilla fruticosa, forces generated by fibroblasts; and Poterium sanguisorba, Pyrus communis, Raphanus (e) selecting an extract that is capable of modulating one raphanistrum, Rheumxhybridum, Rhus typhina L., Ribes or more of said cellular activities to provide a plant nigrum L., Ribes Sylvestre, Rodgersia spp., Rosmarinus extract suitable for the preparation of a dermatological officinalis, Rubus occidentalis, Rubus thibetanus, Rumex formulation. crispus, Rumex scutatus, Ruta graveolens, Salvia officinalis, 19. The process according to claim 18, wherein said Sambucus Canadensis L., Setaria italica, Solanum melon plurality of potential extracts is generated from plant mate gena L., Sorghum dochna bicolor gir technicum, Stellaria rial from a single plant Source. media, Tanacetum cinerariifolium, Taraxacum officinale, US 2007/01 22492 A1 May 31, 2007 76

Teucrium chamaedrys, Thymus fragantissimus, ThymusX 28. The dermatological formulation according to claim 1, citriodorus, Trifolium incarnatum, Triticosecale spp., Tro wherein said plant is subjected to one or more stress prior to paeolum maius L., Tsuga Canadensis, Tsuga diversifolia, said solvent extraction. Vaccinium angustifolium, Vaccinium angustifolium Ait., 29. A method of preparing a dermatological formulation Vitia sp., xTriticosecale spp., Zea mays L. and Zingiber comprising admixing an effective amount of one or more officinale. 26. The dermatological formulation according to claim 1, plant extracts having extracellular protease inhibiting activ wherein said plant extract having extracellular protease ity with a physiologically acceptable carrier, said plant inhibiting activity is derived by solvent extraction from a extract derived from any one of the plants listed in Tables 1, plant selected from the group of Beta vulgaris L., Brassica 2, 3, 4 and 5 by solvent extraction, and said extracellular oleracea L., Capsicum annuum L. Chenopodium quinoa, protease selected from the group of matrix metallopro Daucus carota L., Geraniumxcantabrigiense, Juniperus tease-1 (MMP-1), matrix metalloprotease-2 (MMP-2), communis L., Melilotus alba, Pastinaca sativa L., Potentilla matrix metalloprotease-3 (MMP-3), matrix metallopro anserina L., Rhus typhina L., Solanum melongena L., Tro tease-9 (MMP-9) and human leukocyte elastase (HLE), paeolum maius L., Vaccinium angustifolium, XTriticosecale wherein said plant extract modulates one or more cellular spp. and Zea may’s L. activities in skin cells. 27. The dermatological formulation according to claim 3, wherein said alcoholic solvent is ethanol or a glycol.